Honors & Awards

  • Dean’s Fellowship (Berry award), Stanford (2003-2004)
  • Postdoctoral Fellowship, Lymphoma Research Foundation (2004-2006)

Education & Certifications

  • MSc, King’s College, University of London, Immunology (1995)
  • PhD, King’s College, University of London, Immunogenetics (2002)


All Publications

  • Two alternate strategies for innate immunity to Epstein-Barr virus: One using NK cells and the other NK cells and ?d T cells. journal of experimental medicine Djaoud, Z., Guethlein, L. A., Horowitz, A., Azzi, T., Nemat-Gorgani, N., Olive, D., Nadal, D., Norman, P. J., Münz, C., Parham, P. 2017; 214 (6): 1827-1841


    Most humans become infected with Epstein-Barr virus (EBV), which then persists for life. Infrequently, EBV infection causes infectious mononucleosis (IM) or Burkitt lymphoma (BL). Type I EBV infection, particularly type I BL, stimulates strong responses of innate immune cells. Humans respond to EBV in two alternative ways. Of 24 individuals studied, 13 made strong NK and γδ T cell responses, whereas 11 made feeble γδ T cell responses but stronger NK cell responses. The difference does not correlate with sex, HLA type, or previous exposure to EBV or cytomegalovirus. Cohorts of EBV(+) children and pediatric IM patients include both group 1 individuals, with high numbers of γδ T cells, and group 2 individuals, with low numbers. The even balance of groups 1 and 2 in the human population points to both forms of innate immune response to EBV having benefit for human survival. Correlating these distinctive responses with the progress of EBV infection might facilitate the management of EBV-mediated disease.

    View details for DOI 10.1084/jem.20161017

    View details for PubMedID 28468758

  • Sequences of 95 human MHC haplotypes reveal extreme coding variation in genes other than highly polymorphic HLA class I and II GENOME RESEARCH Norman, P. J., Norberg, S. J., Guethlein, L. A., Nemat-Gorgani, N., Royce, T., Wroblewski, E. E., Dunn, T., Mann, T., Alicata, C., Hollenbach, J. A., Chang, W., Won, M. S., Gunderson, K. L., Abi-Rached, L., Ronaghi, M., Parham, P. 2017; 27 (5): 813-823
  • Distinguishing functional polymorphism from random variation in the sequences of >10,000 HLA-A, -B and -C alleles. PLoS genetics Robinson, J., Guethlein, L. A., Cereb, N., Yang, S. Y., Norman, P. J., Marsh, S. G., Parham, P. 2017; 13 (6): e1006862


    HLA class I glycoproteins contain the functional sites that bind peptide antigens and engage lymphocyte receptors. Recently, clinical application of sequence-based HLA typing has uncovered an unprecedented number of novel HLA class I alleles. Here we define the nature and extent of the variation in 3,489 HLA-A, 4,356 HLA-B and 3,111 HLA-C alleles. This analysis required development of suites of methods, having general applicability, for comparing and analyzing large numbers of homologous sequences. At least three amino-acid substitutions are present at every position in the polymorphic α1 and α2 domains of HLA-A, -B and -C. A minority of positions have an incidence >1% for the 'second' most frequent nucleotide, comprising 70 positions in HLA-A, 85 in HLA-B and 54 in HLA-C. The majority of these positions have three or four alternative nucleotides. These positions were subject to positive selection and correspond to binding sites for peptides and receptors. Most alleles of HLA class I (>80%) are very rare, often identified in one person or family, and they differ by point mutation from older, more common alleles. These alleles with single nucleotide polymorphisms reflect the germ-line mutation rate. Their frequency predicts the human population harbors 8-9 million HLA class I variants. The common alleles of human populations comprise 42 core alleles, which represent all selected polymorphism, and recombinants that have assorted this polymorphism.

    View details for DOI 10.1371/journal.pgen.1006862

    View details for PubMedID 28650991

  • Class I HLA haplotypes form two schools that educate NK cells in different ways. Science immunology Horowitz, A., Djaoud, Z., Nemat-Gorgani, N., Blokhuis, J., Hilton, H. G., Béziat, V., Malmberg, K., Norman, P. J., Guethlein, L. A., Parham, P. 2016; 1 (3)


    Natural killer (NK) cells are lymphocytes having vital functions in innate and adaptive immunity, as well as placental reproduction. Controlling education and functional activity of human NK cells are various receptors that recognize HLA class I on the surface of tissue cells. Epitopes of polymorphic HLA-A,-B and -C are recognized by equally diverse killer cell immunoglobulin-like receptors (KIR). In addition, a peptide cleaved from the leader sequence of HLA-A,-B or -C must bind to HLA-E for it to become a ligand for the conserved CD94:NKG2A receptor. Methionine/threonine dimorphism at position -21 of the leader sequence divides HLA-B allotypes into a majority having -21T that do not supply HLA-E binding peptides and a minority having -21M, that do. Genetic analysis of human populations worldwide shows how haplotypes with -21M HLA-B rarely encode the KIR ligands: Bw4(+)HLA-B and C2(+)HLA-C KIR. Thus there are two fundamental forms of HLA haplotype: one preferentially supplying CD94:NKG2A ligands, the other preferentially supplying KIR ligands. -21 HLA-B dimorphism divides the human population into three groups: M/M, M/T and T/T. Mass cytometry and assays of immune function, shows how M/M and M/T individuals have CD94:NKG2A(+) NK cells which are better educated, phenotypically more diverse and functionally more potent than those in T/T individuals. Fundamental new insights are given to genetic control of NK cell immunity and the evolution that has limited the number of NK cell receptor ligands encoded by an HLA haplotype. These finding suggest new ways to dissect the numerous clinical associations with HLA class I.

    View details for PubMedID 27868107

  • Defining KIR and HLA Class I Genotypes at Highest Resolution via High-Throughput Sequencing. American journal of human genetics Norman, P. J., Hollenbach, J. A., Nemat-Gorgani, N., Marin, W. M., Norberg, S. J., Ashouri, E., Jayaraman, J., Wroblewski, E. E., Trowsdale, J., Rajalingam, R., Oksenberg, J. R., Chiaroni, J., Guethlein, L. A., Traherne, J. A., Ronaghi, M., Parham, P. 2016; 99 (2): 375-391


    The physiological functions of natural killer (NK) cells in human immunity and reproduction depend upon diverse interactions between killer cell immunoglobulin-like receptors (KIRs) and their HLA class I ligands: HLA-A, HLA-B, and HLA-C. The genomic regions containing the KIR and HLA class I genes are unlinked, structurally complex, and highly polymorphic. They are also strongly associated with a wide spectrum of diseases, including infections, autoimmune disorders, cancers, and pregnancy disorders, as well as the efficacy of transplantation and other immunotherapies. To facilitate study of these extraordinary genes, we developed a method that captures, sequences, and analyzes the 13 KIR genes and HLA-A, HLA-B, and HLA-C from genomic DNA. We also devised a bioinformatics pipeline that attributes sequencing reads to specific KIR genes, determines copy number by read depth, and calls high-resolution genotypes for each KIR gene. We validated this method by using DNA from well-characterized cell lines, comparing it to established methods of HLA and KIR genotyping, and determining KIR genotypes from 1000 Genomes sequence data. This identified 116 previously uncharacterized KIR alleles, which were all demonstrated to be authentic by sequencing from source DNA via standard methods. Analysis of just two KIR genes showed that 22% of the 1000 Genomes individuals have a previously uncharacterized allele or a structural variant. The method we describe is suited to the large-scale analyses that are needed for characterizing human populations and defining the precise HLA and KIR factors associated with disease. The methods are applicable to other highly polymorphic genes.

    View details for DOI 10.1016/j.ajhg.2016.06.023

    View details for PubMedID 27486779

  • Co-evolution of Human Leukocyte Antigen (HLA) Class I Ligands with Killer-Cell Immunoglobulin-Like Receptors (KIR) in a Genetically Diverse Population of Sub-Saharan Africans. PLoS genetics Norman, P. J., Hollenbach, J. A., Nemat-Gorgani, N., Guethlein, L. A., Hilton, H. G., Pando, M. J., Koram, K. A., Riley, E. M., Abi-Rached, L., Parham, P. 2013; 9 (10)


    Interactions between HLA class I molecules and killer-cell immunoglobulin-like receptors (KIR) control natural killer cell (NK) functions in immunity and reproduction. Encoded by genes on different chromosomes, these polymorphic ligands and receptors correlate highly with disease resistance and susceptibility. Although studied at low-resolution in many populations, high-resolution analysis of combinatorial diversity of HLA class I and KIR is limited to Asian and Amerindian populations with low genetic diversity. At the other end of the spectrum is the West African population investigated here: we studied 235 individuals, including 104 mother-child pairs, from the Ga-Adangbe of Ghana. This population has a rich diversity of 175 KIR variants forming 208 KIR haplotypes, and 81 HLA-A, -B and -C variants forming 190 HLA class I haplotypes. Each individual we studied has a unique compound genotype of HLA class I and KIR, forming 1-14 functional ligand-receptor interactions. Maintaining this exceptionally high polymorphism is balancing selection. The centromeric region of the KIR locus, encoding HLA-C receptors, is highly diverse whereas the telomeric region encoding Bw4-specific KIR3DL1, lacks diversity in Africans. Present in the Ga-Adangbe are high frequencies of Bw4-bearing HLA-B*53:01 and Bw4-lacking HLA-B*35:01, which otherwise are identical. Balancing selection at key residues maintains numerous HLA-B allotypes having and lacking Bw4, and also those of stronger and weaker interaction with LILRB1, a KIR-related receptor. Correspondingly, there is a balance at key residues of KIR3DL1 that modulate its level of cell-surface expression. Thus, capacity to interact with NK cells synergizes with peptide binding diversity to drive HLA-B allele frequency distribution. These features of KIR and HLA are consistent with ongoing co-evolution and selection imposed by a pathogen endemic to West Africa. Because of the prevalence of malaria in the Ga-Adangbe and previous associations of cerebral malaria with HLA-B*53:01 and KIR, Plasmodium falciparum is a candidate pathogen.

    View details for DOI 10.1371/journal.pgen.1003938

    View details for PubMedID 24204327

  • Bonobos Maintain Immune System Diversity with Three Functional Types of MHC-B JOURNAL OF IMMUNOLOGY Wroblewski, E. E., Guethlein, L. A., Norman, P. J., Li, Y., Shaw, C. M., Han, A. S., Ndjango, J. N., Ahuka-Mundeke, S., Georgiev, A. V., Peeters, M., Hahn, B. H., Parham, P. 2017; 198 (9): 3480-3493


    Fast-evolving MHC class I polymorphism serves to diversify NK cell and CD8 T cell responses in individuals, families, and populations. Because only chimpanzee and bonobo have strict orthologs of all HLA class I, their study gives unique perspectives on the human condition. We defined polymorphism of Papa-B, the bonobo ortholog of HLA-B, for six wild bonobo populations. Sequences for Papa-B exon 2 and 3 were determined from the genomic DNA in 255 fecal samples, minimally representing 110 individuals. Twenty-two Papa-B alleles were defined, each encoding a different Papa-B protein. No Papa-B is identical to any chimpanzee Patr-B, human HLA-B, or gorilla Gogo-B. Phylogenetic analysis identified a clade of MHC-B, defined by residues 45-74 of the α1 domain, which is broadly conserved among bonobo, chimpanzee, and gorilla. Bonobo populations have 3-14 Papa-B allotypes. Three Papa-B are in all populations, and they are each of a different functional type: allotypes having the Bw4 epitope recognized by killer cell Ig-like receptors of NK cells, allotypes having the C1 epitope also recognized by killer cell Ig-like receptors, and allotypes having neither epitope. For population Malebo, these three Papa-B are the only Papa-B allotypes. Although small in number, their sequence divergence is such that the nucleotide diversity (mean proportional distance) of Papa-B in Malebo is greater than in the other populations and is also greater than expected for random combinations of three Papa-B Overall, Papa-B has substantially less diversity than Patr-B in chimpanzee subspecies and HLA-B in indigenous human populations, consistent with bonobo having experienced narrower population bottlenecks.

    View details for DOI 10.4049/jimmunol.601955

    View details for Web of Science ID 000401135200016

    View details for PubMedID 28348269

  • Two Orangutan Species Have Evolved Different KIR Alleles and Haplotypes JOURNAL OF IMMUNOLOGY Guethlein, L. A., Norman, P. J., Heijmans, C. M., de Groot, N. G., Hilton, H. G., Babrzadeh, F., Abi-Rached, L., Bontrop, R. E., Parham, P. 2017; 198 (8): 3157-3169
  • Resurrecting KIR2DP1: A Key Intermediate in the Evolution of Human Inhibitory NK Cell Receptors That Recognize HLA-C. Journal of immunology Hilton, H. G., Blokhuis, J. H., Guethlein, L. A., Norman, P. J., Parham, P. 2017; 198 (5): 1961-1973


    KIR2DP1 is an inactive member of the human lineage III KIR family, which includes all HLA-C-specific receptor genes. The lethal, and only, defect in KIR2DP1 is a nucleotide deletion in codon 88. Fixed in modern humans, the deletion is also in archaic human genomes. KIR2DP1 is polymorphic, with dimorphism at specificity-determining position 44. By repairing the deletion, we resurrected 11 alleles of KIR2DP1(F) , the functional antecedent of KIR2DP1 We demonstrate how K44-KIR2DP1(F) with lysine 44 recognized C1(+)HLA-C, whereas T44-KIR2DP1(F) recognized C2(+)HLA-C. Dimorphisms at 12 other KIR2DP1(F) residues modulate receptor avidity or signaling. KIR2DP1 and KIR2DL1 are neighbors in the centromeric KIR region and are in tight linkage disequilibrium. Like KIR2DL1, KIR2DP1 contributed to CenA and CenB KIR haplotype differences. Encoded on CenA, C1-specific K44-KIR2DP1(F) were stronger receptors than the attenuated C2-specific T44-KIR2DP1(F) encoded on CenB The last common ancestor of humans and chimpanzees had diverse lineage III KIR that passed on to chimpanzees but not to humans. Early humans inherited activating KIR2DS4 and an inhibitory lineage III KIR, likely encoding a C1-specific receptor. The latter spawned the modern family of HLA-C receptors. KIR2DP1(F) has properties consistent with KIR2DP1(F) having been the founder gene. The first KIR2DP1(F) alleles encoded K44-C1 receptors; subsequently KIR2DP1(F) alleles encoding T44-C2 receptors evolved. The emergence of dedicated KIR2DL2/3 and KIR2DL1 genes encoding C1 and C2 receptors, respectively, could have led to obsolescence of KIR2DP1(F) Alternatively, pathogen subversion caused its demise. Preservation of KIR2DP1(F) functional polymorphism was a side effect of fixation of the deletion in KIR2DP1(F) by micro gene conversion.

    View details for DOI 10.4049/jimmunol.1601835

    View details for PubMedID 28122963

    View details for PubMedCentralID PMC5321844

  • Deciphering the killer-cell immunoglobulin-like receptor system at super-resolution for natural killer and T-cell biology IMMUNOLOGY Beziat, V., Hilton, H. G., Norman, P. J., Traherne, J. A. 2017; 150 (3): 248-264

    View details for DOI 10.1111/imm.12684

    View details for Web of Science ID 000394789800002

  • KIR2DS5 allotypes that recognize the C2 epitope of HLA-C are common among Africans and absent from Europeans. Immunity, inflammation and disease Blokhuis, J. H., Hilton, H. G., Guethlein, L. A., Norman, P. J., Nemat-Gorgani, N., Nakimuli, A., Chazara, O., Moffett, A., Parham, P. 2017


    KIR2DS5 is an activating human NK cell receptor of lineage III KIR. These include both inhibitory KIR2DL1, 2 and 3 and activating KIR2DS1 that recognize either the C1 or C2 epitope of HLA-C. In Europeans KIR2DS5 is essentially monomorphic, with KIR2DS5*002 being predominant. Pioneering investigations showed that KIR2DS5*002 has activating potential, but cannot recognize HLA-A, -B, or -C. Subsequent studies have shown that KIR2DS5 is highly polymorphic in Africans, and that KIR2DS5*006 protects pregnant Ugandan women from preeclampsia. Because inhibitory C2-specific KIR2DL1 correlates with preeclampsia, whereas activating C2-specific KIR2DS1 protects, this association pointed to KIR2DS5*006 being an activating C2-specific receptor. To test this hypothesis we made KIR-Fc fusion proteins from all ten KIR2DS5 allotypes and tested their binding to a representative set of HLA-A, -B and -C allotypes.Six African-specific KIR2DS5 bound to C2(+) HLA-C but not to other HLA class I. Their avidity for C2 is ∼20% that of C2-specific KIR2DL1 and ∼40% that of C2-specific KIR2DS1. Among the African C2 receptors is KIR2DS5*006, which protected a cohort of pregnant Ugandans from pre-eclampsia. Three African KIR2DS5 allotypes and KIR2DS5*002, bound no HLA-A, -B or -C. As a group the C2-binding KIR2DS5 allotypes protect against pre-eclampsia compared to the non-binding KIR2DS5 allotypes. Natural substitutions that contribute to loss or reduction of C2 receptor function are at positions 127, 158, and 176 in the D2 domain.KIR2DS5*005 has the KIR2DS5 consensus sequence, is the only allele found at both centromeric and telomeric locations of KIR2DS5, and is likely the common ancestor of all KIR2DS5 alleles. That KIR2DS5*005 has C2 receptor activity, points to KIR2DS5*002, and other allotypes lacking C2 receptor function, being products of attenuation, a characteristic feature of most KIR B haplotype genes. Alleles encoding attenuated and active KIR2DS5 are present in both centromeric and telomeric locations.

    View details for DOI 10.1002/iid3.178

    View details for PubMedID 28685972

  • HLA class I variation in Iranian Lur and Kurd populations: high haplotype and allotype diversity with an abundance of KIR ligands. HLA Ashouri, E., Norman, P. J., Guethlein, L. A., Han, A. S., Nemat-Gorgani, N., Norberg, S. J., Ghaderi, A., Parham, P. 2016; 88 (3): 87-99


    HLA-A, -B and -C alleles of 285 individuals, representing three Iranian Lur populations and one Iranian Kurd population were sequenced completely, yielding human leukocyte antigen (HLA) class I genotypes at high resolution and filling four fields of the official HLA nomenclature. Each population has 87-99 alleles, evenly distributed between the three HLA class I genes, 145 alleles being identified in total. These alleles were already known, named and deposited in the HLA database. The alleles form 316 different HLA A-B-C haplotypes, with each population having between 80 and 112 haplotypes. The four Iranian populations form a related group that is distinguished from other populations, including other Iranians. All four KIR ligands - the A3/11, Bw4, C1 and C2 epitopes - are well represented, particularly Bw4, which is carried by three high-frequency allotypes: HLA-A*24:02, HLA-A*32:01 and HLA-B*51:01. In the Lur and Kurd populations, between 82% and 94% of individuals have the Bw4 epitope, the ligand for KIR3DL1. HLA-B*51:01 is likely of Neandertal origin and associated with Behcet's disease, also known as the Silk Road disease. The Lordegan Lur have the highest frequency of HLA-B*51:01 in the world. This allele is present on 46 Lur and Kurd haplotypes. Present at lower frequency is HLA-B*51:08, which is also associated with Behcet's disease. In the four Iranian populations, 31 haplotypes encode both Bw4(+) HLA-A and Bw4(+) HLA-B, a dual combination of Bw4 epitopes that is relatively rare in other populations, worldwide. This study both demonstrates and emphasizes the value of studying HLA class I polymorphism at highest resolution in anthropologically well-defined populations.

    View details for DOI 10.1111/tan.12852

    View details for PubMedID 27558013

  • Hematopoietic stem cell transplantation: Improving alloreactive Bw4 donor selection by genotyping codon 86 of KIR3DL1/S1. European journal of immunology Alicata, C., Pende, D., Meazza, R., Canevali, P., Loiacono, F., Bertaina, A., Locatelli, F., Nemat-Gorgani, N., Guethlein, L. A., Parham, P., Moretta, L., Moretta, A., Bottino, C., Norman, P. J., Falco, M. 2016; 46 (6): 1511-1517


    KIR3DL1 is a natural killer (NK) cell receptor that recognizes the Bw4 epitope of human leukocyte antigen (HLA) class I molecules. Following hematopoietic stem cell transplantation for patients lacking Bw4, KIR3DL1-expressing NK cells from Bw4-positive donors can be alloreactive and eliminate tumor cells. However, KIR3DL1 alleles having T instead of C at nucleotide 320 (encoding leucine 86 instead of serine 86) are not expressed on the cell surface. Thus, not all individuals testing positive for KIR3DL1 are optimal donors for Bw4-negative recipients. Therefore, we developed a method for genotyping codon 86, which was validated by its perfect correlation with NK cell phenotype for 100 donors of diverse KIR3DL1/S1 genotype. We typed 600 donors and found that ∼12.2% had the KIR3DL1 gene, but did not express cell-surface KIR3DL1. By contrast, high-expressing allotypes were identified when haplotypes from four families with duplicated KIR3DL1/S1 genes were characterized at high resolution. Identifying donors who have KIR3DL1 but lack cell-surface KIR3DL1 would refine donor selection. With this technique, the number of individuals identified who may not be optimal donors for Bw4-negative patients increases by threefold, when compared with standard methods. Taken together, we propose that allele typing of killer cell Ig-like receptor (KIR) polymorphisms should become a standard practice when selecting donors.

    View details for DOI 10.1002/eji.201546236

    View details for PubMedID 26990677

  • Description of the novel KIR2DL4*035 allele identified using high-throughput sequencing. HLA : immune response genetics Alicata, C., Bottino, C., Guethlein, L. A., Parham, P., Norman, P. J. 2016; 87 (3): 191-193


    A newly identified allele of the KIR2DL4 natural killer cell receptor for human leukocyte antigen (HLA) class I.

    View details for DOI 10.1111/tan.12761

    View details for PubMedID 26917249

  • Minimum information for reporting next generation sequence genotyping (MIRING): Guidelines for reporting HLA and KIR genotyping via next generation sequencing HUMAN IMMUNOLOGY Mack, S. J., Milius, R. P., Gifford, B. D., Sauter, J., Hofmann, J., Osoegawa, K., Robinson, J., Groeneweg, M., Turenchalk, G. S., Adai, A., Holcomb, C., Rozemuller, E. H., Penning, M. T., Heuer, M. L., Wang, C., Salit, M. L., Schmidt, A. H., Parham, P. R., Mueller, C., Hague, T., Fischer, G., Fernandez-Vina, M., Hollenbach, J. A., Norman, P. J., Maiers, M. 2015; 76 (12): 954-962

    View details for DOI 10.1016/j.humimm.2015.09.011

    View details for Web of Science ID 000366437900012

    View details for PubMedID 26407912

  • Regulation of Adaptive NK Cells and CD8 T Cells by HLA-C Correlates with Allogeneic Hematopoietic Cell Transplantation and with Cytomegalovirus Reactivation JOURNAL OF IMMUNOLOGY Horowitz, A., Guethlein, L. A., Nemat-Gorgani, N., Norman, P. J., Cooley, S., Miller, J. S., Parham, P. 2015; 195 (9): 4524-4536


    Mass cytometry was used to investigate the effect of CMV reactivation on lymphocyte reconstitution in hematopoietic cell transplant patients. For eight transplant recipients (four CMV negative and four CMV positive), we studied PBMCs obtained 6 mo after unrelated donor hematopoietic cell transplantation (HCT). Forty cell-surface markers, distinguishing all major leukocyte populations in PBMC, were analyzed with mass cytometry. This group included 34 NK cell markers. Compared with healthy controls, transplant recipients had higher HLA-C expression on CD56(-)CD16(+) NK cells, B cells, CD33(bright) myeloid cells, and CD4CD8 T cells. The increase in HLA-C expression was greater for CMV-positive HCT recipients than for CMV negative recipients. Present in CMV-positive HCT recipients, but not in CMV-negative HCT recipients or controls, is a population of killer cell Ig-like receptor (KIR)-expressing CD8 T cells not previously described. These CD8 T cells coexpress CD56, CD57, and NKG2C. The HCT recipients also have a population of CD57(+)NKG2A(+) NK cells that preferentially express KIR2DL1. An inverse correlation was observed between the frequencies of CD57(+)NKG2C(+) NK cells and CD57(+)NKG2A(+) NK cells. Although CD57(+)NKG2A(+) NK cells are less abundant in CMV-positive recipients, their phenotype is of a more activated cell than the CD57(+)NKG2A(+) NK cells of controls and CMV-negative HCT recipients. These data demonstrate that HCT and CMV reactivation are associated with an increased expression of HLA-C. This could influence NK cell education during lymphocyte reconstitution. The increased inhibitory KIR expression by proliferating CMV-specific CD8 T cells suggests regulatory interactions between HLA-C and KIR might promote Graft-versus-Leukemia effects following transplantation.

    View details for DOI 10.4049/jimmunol.1401990

    View details for Web of Science ID 000362968500049

    View details for PubMedID 26416275

  • The production of KIR-Fc fusion proteins and their use in a multiplex HLA class I binding assay. Journal of immunological methods Hilton, H. G., Moesta, A. K., Guethlein, L. A., Blokhuis, J., Parham, P., Norman, P. J. 2015; 425: 79-87


    Soluble recombinant proteins that comprise the extracellular part of a surface expressed receptor attached to the Fc region of an IgG antibody have facilitated the determination of ligand specificity for an array of immune system receptors. Among such receptors is the family of killer cell immunoglobulin-like receptors (KIR) that recognize HLA class I ligands. These receptors, expressed on natural killer (NK) cells and T cells, play important roles in both immune defense and placental development in early pregnancy. Here we describe a method for the production of two domain KIR-Fc fusion proteins using baculovirus infected insect cells. This method is more scalable than traditional mammalian cell expression systems and produces efficiently folded proteins that carry posttranslational modifications found in native KIR. We also describe a multiplex binding assay using the Luminex platform that determines the avidity and specificity of two domain KIR-Fc for a panel of microbeads, each coated with one of 97 HLA class I allotypes. This assay is simple to perform, and represents a major improvement over the assays used previously, which were limited in the number of KIR and HLA class I combinations that could be assayed at any one time. The results obtained from this assay can be used to predict the response of NK cell and T cells when their KIR recognize HLA class I.

    View details for DOI 10.1016/j.jim.2015.06.012

    View details for PubMedID 26096968

  • Polymorphic HLA-C Receptors Balance the Functional Characteristics of KIR Haplotypes. Journal of immunology Hilton, H. G., Guethlein, L. A., Goyos, A., Nemat-Gorgani, N., Bushnell, D. A., Norman, P. J., Parham, P. 2015; 195 (7): 3160-3170


    The human killer cell Ig-like receptor (KIR) locus comprises two groups of KIR haplotypes, termed A and B. These are present in all human populations but with different relative frequencies, suggesting they have different functional properties that underlie their balancing selection. We studied the genomic organization and functional properties of the alleles of the inhibitory and activating HLA-C receptors encoded by KIR haplotypes. Because every HLA-C allotype functions as a ligand for KIR, the interactions between KIR and HLA-C dominate the HLA class I-mediated regulation of human NK cells. The C2 epitope is recognized by inhibitory KIR2DL1 and activating KIR2DS1, whereas the C1 epitope is recognized by inhibitory KIR2DL2 and KIR2DL3. This study shows that the KIR2DL1, KIR2DS1, and KIR2DL2/3 alleles form distinctive phylogenetic clades that associate with specific KIR haplotypes. KIR A haplotypes are characterized by KIR2DL1 alleles that encode strong inhibitory C2 receptors and KIR2DL3 alleles encoding weak inhibitory C1 receptors. In striking contrast, KIR B haplotypes are characterized by KIR2DL1 alleles that encode weak inhibitory C2 receptors and KIR2DL2 alleles encoding strong inhibitory C1 receptors. The wide-ranging properties of KIR allotypes arise from substitutions throughout the KIR molecule. Such substitutions can influence cell surface expression, as well as the avidity and specificity for HLA-C ligands. Consistent with the crucial role of inhibitory HLA-C receptors in self-recognition, as well as NK cell education and response, most KIR haplotypes have both a functional C1 and C2 receptor, despite the considerable variation that occurs in ligand recognition and surface expression.

    View details for DOI 10.4049/jimmunol.1501358

    View details for PubMedID 26311903

  • Very long haplotype tracts characterized at high resolution from HLA homozygous cell lines IMMUNOGENETICS Norman, P. J., Norberg, S. J., Nemat-Gorgani, N., Royce, T., Hollenbach, J. A., Won, M. S., Guethlein, L. A., Gunderson, K. L., Ronaghi, M., Parham, P. 2015; 67 (9): 479-485
  • Co-evolution of MHC class I and variable NK cell receptors in placental mammals IMMUNOLOGICAL REVIEWS Guethlein, L. A., Norman, P. J., Hilton, H. H., Parham, P. 2015; 267 (1): 259-282

    View details for DOI 10.1111/imr.12326

    View details for Web of Science ID 000360082000017

  • Race, Ethnicity and Ancestry in Unrelated Transplant Matching for the National Marrow Donor Program: A Comparison of Multiple Forms of Self-Identification with Genetics PLOS ONE Hollenbach, J. A., Saperstein, A., Albrecht, M., Vierra-Green, C., Parham, P., Norman, P. J., Maiers, M. 2015; 10 (8)
  • Loss and Gain of Natural Killer Cell Receptor Function in an African Hunter-Gatherer Population. PLoS genetics Hilton, H. G., Norman, P. J., Nemat-Gorgani, N., Goyos, A., Hollenbach, J. A., Henn, B. M., Gignoux, C. R., Guethlein, L. A., Parham, P. 2015; 11 (8)


    Modulating natural killer cell functions in human immunity and reproduction are diverse interactions between the killer cell immunoglobulin-like receptors (KIR) of Natural Killer (NK) cells and HLA class I ligands on the surface of tissue cells. Dominant interactions are between KIR2DL1 and the C2 epitope of HLA-C and between KIR2DL2/3 and the C1 epitope of HLA-C. KhoeSan hunter-gatherers of Southern Africa represent the earliest population divergence known and are the most genetically diverse indigenous people, qualities reflected in their KIR and HLA genes. Of the ten KhoeSan KIR2DL1 alleles, KIR2DL1*022 and KIR2DL1*026 likely originated in the KhoeSan, and later were transmitted at low frequency to the neighboring Zulus through gene flow. These alleles arose by point mutation from other KhoeSan KIR2DL1 alleles that are more widespread globally. Mutation of KIR2DL1*001 gave rise to KIR2DL1*022, causing loss of C2 recognition and gain of C1 recognition. This makes KIR2DL1*022 a more avid and specific C1 receptor than any KIR2DL2/3 allotype. Mutation of KIR2DL1*012 gave rise to KIR2DL1*026, causing premature termination of translation at the end of the transmembrane domain. This makes KIR2DL1*026 a membrane-associated receptor that lacks both a cytoplasmic tail and signaling function. At higher frequencies than their parental allotypes, the combined effect of the KhoeSan-specific KIR2DL1*022 and KIR2DL1*026 is to reduce the frequency of strong inhibitory C2 receptors and increase the frequency of strong inhibitory C1 receptors. Because interaction of KIR2DL1 with C2 is associated with risk of pregnancy disorder, these functional changes are potentially advantageous. Whereas all other KhoeSan KIR2DL1 alleles are present on a wide diversity of centromeric KIR haplotypes, KIR2DL1*026 is present on a single KIR haplotype and KIR2DL1*022 is present on two very similar haplotypes. The high linkage disequilibrium across their haplotypes is consistent with a recent emergence for these KIR2DL1 alleles that have distinctive functions.

    View details for DOI 10.1371/journal.pgen.1005439

    View details for PubMedID 26292085

    View details for PubMedCentralID PMC4546388

  • Signature Patterns of MHC Diversity in Three Gombe Communities of Wild Chimpanzees Reflect Fitness in Reproduction and Immune Defense against SIVcpz. PLoS biology Wroblewski, E. E., Norman, P. J., Guethlein, L. A., Rudicell, R. S., Ramirez, M. A., Li, Y., Hahn, B. H., Pusey, A. E., Parham, P. 2015; 13 (5)


    Major histocompatibility complex (MHC) class I molecules determine immune responses to viral infections. These polymorphic cell-surface glycoproteins bind peptide antigens, forming ligands for cytotoxic T and natural killer cell receptors. Under pressure from rapidly evolving viruses, hominoid MHC class I molecules also evolve rapidly, becoming diverse and species-specific. Little is known of the impact of infectious disease epidemics on MHC class I variant distributions in human populations, a context in which the chimpanzee is the superior animal model. Population dynamics of the chimpanzees inhabiting Gombe National Park, Tanzania have been studied for over 50 years. This population is infected with SIVcpz, the precursor of human HIV-1. Because HLA-B is the most polymorphic human MHC class I molecule and correlates strongly with HIV-1 progression, we determined sequences for its ortholog, Patr-B, in 125 Gombe chimpanzees. Eleven Patr-B variants were defined, as were their frequencies in Gombe's three communities, changes in frequency with time, and effect of SIVcpz infection. The growing populations of the northern and central communities, where SIVcpz is less prevalent, have stable distributions comprising a majority of low-frequency Patr-B variants and a few high-frequency variants. Driving the latter to high frequency has been the fecundity of immigrants to the northern community, whereas in the central community, it has been the fecundity of socially dominant individuals. In the declining population of the southern community, where greater SIVcpz prevalence is associated with mortality and emigration, Patr-B variant distributions have been changing. Enriched in this community are Patr-B variants that engage with natural killer cell receptors. Elevated among SIVcpz-infected chimpanzees, the Patr-B*06:03 variant has striking structural and functional similarities to HLA-B*57, the human allotype most strongly associated with delayed HIV-1 progression. Like HLA-B*57, Patr-B*06:03 correlates with reduced viral load, as assessed by detection of SIVcpz RNA in feces.

    View details for DOI 10.1371/journal.pbio.1002144

    View details for PubMedID 26020813

  • A KIR B centromeric region present in Africans but not Europeans protects pregnant women from pre-eclampsia. Proceedings of the National Academy of Sciences of the United States of America Nakimuli, A., Chazara, O., Hiby, S. E., Farrell, L., Tukwasibwe, S., Jayaraman, J., Traherne, J. A., Trowsdale, J., Colucci, F., Lougee, E., Vaughan, R. W., Elliott, A. M., Byamugisha, J., Kaleebu, P., Mirembe, F., Nemat-Gorgani, N., Parham, P., Norman, P. J., Moffett, A. 2015; 112 (3): 845-850


    In sub-Saharan Africans, maternal mortality is unacceptably high, with >400 deaths per 100,000 births compared with <10 deaths per 100,000 births in Europeans. One-third of the deaths are caused by pre-eclampsia, a syndrome arising from defective placentation. Controlling placentation are maternal natural killer (NK) cells that use killer-cell immunoglobulin-like receptor (KIR) to recognize the fetal HLA-C molecules on invading trophoblast. We analyzed genetic polymorphisms of maternal KIR and fetal HLA-C in 484 normal and 254 pre-eclamptic pregnancies at Mulago Hospital, Kampala, Uganda. The combination of maternal KIR AA genotypes and fetal HLA-C alleles encoding the C2 epitope associates with pre-eclampsia [P = 0.0318, odds ratio (OR) = 1.49]. The KIR genes associated with protection are located in centromeric KIR B regions that are unique to sub-Saharan African populations and contain the KIR2DS5 and KIR2DL1 genes (P = 0.0095, OR = 0.59). By contrast, telomeric KIR B genes protect Europeans against pre-eclampsia. Thus, different KIR B regions protect sub-Saharan Africans and Europeans from pre-eclampsia, whereas in both populations, the KIR AA genotype is a risk factor for the syndrome. These results emphasize the importance of undertaking genetic studies of pregnancy disorders in African populations with the potential to provide biological insights not available from studies restricted to European populations.

    View details for DOI 10.1073/pnas.1413453112

    View details for PubMedID 25561558

  • POPULATION GENETICS. Genomic evidence for the Pleistocene and recent population history of Native Americans. Science (New York, N.Y.) Raghavan, M., Steinrücken, M., Harris, K., Schiffels, S., Rasmussen, S., DeGiorgio, M., Albrechtsen, A., Valdiosera, C., Ávila-Arcos, M. C., Malaspinas, A. S., Eriksson, A., Moltke, I., Metspalu, M., Homburger, J. R., Wall, J., Cornejo, O. E., Moreno-Mayar, J. V., Korneliussen, T. S., Pierre, T., Rasmussen, M., Campos, P. F., Damgaard, P. d., Allentoft, M. E., Lindo, J., Metspalu, E., Rodríguez-Varela, R., Mansilla, J., Henrickson, C., Seguin-Orlando, A., Malmström, H., Stafford, T., Shringarpure, S. S., Moreno-Estrada, A., Karmin, M., Tambets, K., Bergström, A., Xue, Y., Warmuth, V., Friend, A. D., Singarayer, J., Valdes, P., Balloux, F., Leboreiro, I., Vera, J. L., Rangel-Villalobos, H., Pettener, D., Luiselli, D., Davis, L. G., Heyer, E., Zollikofer, C. P., Ponce de León, M. S., Smith, C. I., Grimes, V., Pike, K. A., Deal, M., Fuller, B. T., Arriaza, B., Standen, V., Luz, M. F., Ricaut, F., Guidon, N., Osipova, L., Voevoda, M. I., Posukh, O. L., Balanovsky, O., Lavryashina, M., Bogunov, Y., Khusnutdinova, E., Gubina, M., Balanovska, E., Fedorova, S., Litvinov, S., Malyarchuk, B., Derenko, M., Mosher, M. J., Archer, D., Cybulski, J., Petzelt, B., Mitchell, J., Worl, R., Norman, P. J., Parham, P., Kemp, B. M., Kivisild, T., Tyler-Smith, C., Sandhu, M. S., Crawford, M., Villems, R., Smith, D. G., Waters, M. R., Goebel, T., Johnson, J. R., Malhi, R. S., Jakobsson, M., Meltzer, D. J., Manica, A., Durbin, R., Bustamante, C. D., Song, Y. S., Nielsen, R., Willerslev, E. 2015; 349 (6250): aab3884


    How and when the Americas were populated remains contentious. Using ancient and modern genome-wide data, we found that the ancestors of all present-day Native Americans, including Athabascans and Amerindians, entered the Americas as a single migration wave from Siberia no earlier than 23 thousand years ago (ka) and after no more than an 8000-year isolation period in Beringia. After their arrival to the Americas, ancestral Native Americans diversified into two basal genetic branches around 13 ka, one that is now dispersed across North and South America and the other restricted to North America. Subsequent gene flow resulted in some Native Americans sharing ancestry with present-day East Asians (including Siberians) and, more distantly, Australo-Melanesians. Putative "Paleoamerican" relict populations, including the historical Mexican Pericúes and South American Fuego-Patagonians, are not directly related to modern Australo-Melanesians as suggested by the Paleoamerican Model.

    View details for DOI 10.1126/science.aab3884

    View details for PubMedID 26198033

    View details for PubMedCentralID PMC4733658

  • Definition of the Cattle Killer Cell Ig-like Receptor Gene Family: Comparison with Aurochs and Human Counterparts JOURNAL OF IMMUNOLOGY Sanderson, N. D., Norman, P. J., Guethlein, L. A., Ellis, S. A., Williams, C., Breen, M., Park, S. D., Magee, D. A., Babrzadeh, F., Warry, A., Watson, M., Bradley, D. G., MacHugh, D. E., Parham, P., Hammond, J. A. 2014; 193 (12): 6016-6030


    Under selection pressure from pathogens, variable NK cell receptors that recognize polymorphic MHC class I evolved convergently in different species of placental mammal. Unexpectedly, diversified killer cell Ig-like receptors (KIRs) are shared by simian primates, including humans, and cattle, but not by other species. Whereas much is known of human KIR genetics and genomics, knowledge of cattle KIR is limited to nine cDNA sequences. To facilitate comparison of the cattle and human KIR gene families, we determined the genomic location, structure, and sequence of two cattle KIR haplotypes and defined KIR sequences of aurochs, the extinct wild ancestor of domestic cattle. Larger than its human counterpart, the cattle KIR locus evolved through successive duplications of a block containing ancestral KIR3DL and KIR3DX genes that existed before placental mammals. Comparison of two cattle KIR haplotypes and aurochs KIR show the KIR are polymorphic and the gene organization and content appear conserved. Of 18 genes, 8 are functional and 10 were inactivated by point mutation. Selective inactivation of KIR3DL and activating receptor genes leaves a functional cohort of one inhibitory KIR3DL, one activating KIR3DX, and six inhibitory KIR3DX. Functional KIR diversity evolved from KIR3DX in cattle and from KIR3DL in simian primates. Although independently evolved, cattle and human KIR gene families share important function-related properties, indicating that cattle KIR are NK cell receptors for cattle MHC class I. Combinations of KIR and MHC class I are the major genetic factors associated with human disease and merit investigation in cattle.

    View details for DOI 10.4049/jimmunol.1401980

    View details for Web of Science ID 000346082400029

    View details for PubMedID 25398326

  • KIR diversity in MAori and Polynesians: populations in which HLA-B is not a significant KIR ligand IMMUNOGENETICS Nemat-Gorgani, N., Edinur, H. A., Hollenbach, J. A., Traherne, J. A., Dunn, P. P., Chambers, G. K., Parham, P., Norman, P. J. 2014; 66 (11): 597-611
  • Exome capture from saliva produces high quality genomic and metagenomic data. BMC genomics Kidd, J. M., Sharpton, T. J., Bobo, D., Norman, P. J., Martin, A. R., Carpenter, M. L., Sikora, M., Gignoux, C. R., Nemat-Gorgani, N., Adams, A., Guadalupe, M., Guo, X., Feng, Q., Li, Y., Liu, X., Parham, P., Hoal, E. G., Feldman, M. W., Pollard, K. S., Wall, J. D., Bustamante, C. D., Henn, B. M. 2014; 15 (1): 262-?

    View details for DOI 10.1186/1471-2164-15-262

    View details for PubMedID 24708091

  • Reconstructing the population genetic history of the Caribbean. PLoS genetics Moreno-Estrada, A., Gravel, S., Zakharia, F., McCauley, J. L., Byrnes, J. K., Gignoux, C. R., Ortiz-Tello, P. A., Martínez, R. J., Hedges, D. J., Morris, R. W., Eng, C., Sandoval, K., Acevedo-Acevedo, S., Norman, P. J., Layrisse, Z., Parham, P., Martínez-Cruzado, J. C., Burchard, E. G., Cuccaro, M. L., Martin, E. R., Bustamante, C. D. 2013; 9 (11)

    View details for DOI 10.1371/journal.pgen.1003925

    View details for PubMedID 24244192

  • Genetic and environmental determinants of human NK cell diversity revealed by mass cytometry. Science translational medicine Horowitz, A., Strauss-Albee, D. M., Leipold, M., Kubo, J., Nemat-Gorgani, N., Dogan, O. C., Dekker, C. L., Mackey, S., Maecker, H., Swan, G. E., Davis, M. M., Norman, P. J., Guethlein, L. A., Desai, M., Parham, P., Blish, C. A. 2013; 5 (208): 208ra145-?

    View details for DOI 10.1126/scitranslmed.3006702

    View details for PubMedID 24154599

  • 16(th) IHIW: Review of HLA typing by NGS INTERNATIONAL JOURNAL OF IMMUNOGENETICS De Santis, D., Dinauer, D., Duke, J., Erlich, H. A., HOLCOMB, C. L., Lind, C., Mackiewicz, K., Monos, D., Moudgil, A., Norman, P., Parham, P., Sasson, A., Allcock, R. J. 2013; 40 (1): 72-76


    Human leucocyte antigen (HLA) genes play an important role in the success of organ transplantation and are associated with autoimmune and infectious diseases. Current DNA-based genotyping methods, including Sanger sequence-based typing (SSBT), have identified a high degree of polymorphism. This level of polymorphism makes high-resolution HLA genotyping challenging, resulting in ambiguous typing results due to an inability to resolve phase and/or defining polymorphisms lying outside the region amplified. Next-generation sequencing (NGS) may resolve the issue through the combination of clonal amplification, which provides phase information, and the ability to sequence larger regions of genes, including introns, without the additional effort or cost associated with current methods. The NGS HLA sequencing project of the 16IHIW aimed to discuss the different approaches to (i) template preparation including short- and long-range PCR amplicons, exome capture and whole genome; (ii) sequencing platforms, including GS 454 FLX, Ion Torrent PGM, Illumina MiSeq/HiSeq and Pacific Biosciences SMRT; (iii) data analysis, specifically allele-calling software. The pilot studies presented at the workshop demonstrated that although individual sequencers have very different performance characteristics, all produced sequence data suitable for the resolution of HLA genotyping ambiguities. The developments presented at this workshop clearly highlight the potential benefits of NGS in the HLA laboratory.

    View details for DOI 10.1111/iji.12024

    View details for Web of Science ID 000313488000012

    View details for PubMedID 23302098

  • Natural selection on marine carnivores elaborated a diverse family of classical MHC class I genes exhibiting haplotypic gene content variation and allelic polymorphism IMMUNOGENETICS Hammond, J. A., Guethlein, L. A., Norman, P. J., Parham, P. 2012; 64 (12): 915-933


    Pinnipeds, marine carnivores, diverged from terrestrial carnivores ~45 million years ago, before their adaptation to marine environments. This lifestyle change exposed pinnipeds to different microbiota and pathogens, with probable impact on their MHC class I genes. Investigating this question, genomic sequences were determined for 71 MHC class I variants: 27 from harbor seal and 44 from gray seal. These variants form three MHC class I gene lineages, one comprising a pseudogene. The second, a candidate nonclassical MHC class I gene, comprises a nonpolymorphic transcribed gene related to dog DLA-79 and giant panda Aime-1906. The third is the diversity lineage, which includes 62 of the 71 seal MHC class I variants. All are transcribed, and they minimally represent six harbor and 12 gray seal MHC class I genes. Besides species-specific differences in gene number, seal MHC class I haplotypes exhibit gene content variation and allelic polymorphism. Patterns of sequence variation, and of positions for positively selected sites, indicate the diversity lineage genes are the seals' classical MHC class I genes. Evidence that expansion of diversity lineage genes began before gray and harbor seals diverged is the presence in both species of two distinctive sublineages of diversity lineage genes. Pointing to further expansion following the divergence are the presence of species-specific genes and greater MHC class I diversity in gray seals than harbor seals. The elaboration of a complex variable family of classical MHC class I genes in pinnipeds contrasts with the single, highly polymorphic classical MHC class I gene of dog and giant panda, terrestrial carnivores.

    View details for DOI 10.1007/s00251-012-0651-z

    View details for Web of Science ID 000311025500007

    View details for PubMedID 23001684

  • Mutation at positively selected positions in the binding site for HLA-C shows that KIR2DL1 is a more refined but less adaptable NK cell receptor than KIR2DL3. Journal of immunology Hilton, H. G., Vago, L., Older Aguilar, A. M., Moesta, A. K., Graef, T., Abi-Rached, L., Norman, P. J., Guethlein, L. A., Fleischhauer, K., Parham, P. 2012; 189 (3): 1418-1430


    Through recognition of HLA class I, killer cell Ig-like receptors (KIR) modulate NK cell functions in human immunity and reproduction. Although a minority of HLA-A and -B allotypes are KIR ligands, HLA-C allotypes dominate this regulation, because they all carry either the C1 epitope recognized by KIR2DL2/3 or the C2 epitope recognized by KIR2DL1. The C1 epitope and C1-specific KIR evolved first, followed several million years later by the C2 epitope and C2-specific KIR. Strong, varying selection pressure on NK cell functions drove the diversification and divergence of hominid KIR, with six positions in the HLA class I binding site of KIR being targets for positive diversifying selection. Introducing each naturally occurring residue at these positions into KIR2DL1 and KIR2DL3 produced 38 point mutants that were tested for binding to 95 HLA- A, -B, and -C allotypes. Modulating specificity for HLA-C is position 44, whereas positions 71 and 131 control cross-reactivity with HLA-A*11:02. Dominating avidity modulation is position 70, with lesser contributions from positions 68 and 182. KIR2DL3 has lower avidity and broader specificity than KIR2DL1. Mutation could increase the avidity and change the specificity of KIR2DL3, whereas KIR2DL1 specificity was resistant to mutation, and its avidity could only be lowered. The contrasting inflexibility of KIR2DL1 and adaptability of KIR2DL3 fit with C2-specific KIR having evolved from C1-specific KIR, and not vice versa. Substitutions restricted to activating KIR all reduced the avidity of KIR2DL1 and KIR2DL3, further evidence that activating KIR function often becomes subject to selective attenuation.

    View details for DOI 10.4049/jimmunol.1100431

    View details for PubMedID 22772445

  • Mutation at Positively Selected Positions in the Binding Site for HLA-C Shows That KIR2DL1 Is a More Refined but Less Adaptable NK Cell Receptor Than KIR2DL3 JOURNAL OF IMMUNOLOGY Hilton, H. G., Vago, L., Aguilar, A. M., Moesta, A. K., Graef, T., Abi-Rached, L., Norman, P. J., Guethlein, L. A., Fleischhauer, K., Parham, P. 2012; 189 (3): 1418-1430
  • Human-specific evolution of killer cell immunoglobulin-like receptor recognition of major histocompatibility complex class I molecules PHILOSOPHICAL TRANSACTIONS OF THE ROYAL SOCIETY B-BIOLOGICAL SCIENCES Parham, P., Norman, P. J., Abi-Rached, L., Guethlein, L. A. 2012; 367 (1590): 800-811


    In placental mammals, natural killer (NK) cells are a population of lymphocytes that make unique contributions to immune defence and reproduction, functions essential for survival of individuals, populations and species. Modulating these functions are conserved and variable NK-cell receptors that recognize epitopes of major histocompatibility complex (MHC) class I molecules. In humans, for example, recognition of human leucocyte antigen (HLA)-E by the CD94:NKG2A receptor is conserved, whereas recognition of HLA-A, B and C by the killer cell immunoglobulin-like receptors (KIRs) is diversified. Competing demands of the immune and reproductive systems, and of T-cell and NK-cell immunity-combined with the segregation on different chromosomes of variable NK-cell receptors and their MHC class I ligands-drive an unusually rapid evolution that has resulted in unprecedented levels of species specificity, as first appreciated from comparison of mice and humans. Counterparts to human KIR are present only in simian primates. Observed in these species is the coevolution of KIR and the four MHC class I epitopes to which human KIR recognition is restricted. Unique to hominids is the emergence of the MHC-C locus as a supplier of specialized and superior ligands for KIR. This evolutionary trend is most highly elaborated in the chimpanzee. Unique to the human KIR locus are two groups of KIR haplotypes that are present in all human populations and subject to balancing selection. Group A KIR haplotypes resemble chimpanzee KIR haplotypes and are enriched for genes encoding KIR that bind HLA class I, whereas group B KIR haplotypes are enriched for genes encoding receptors with diminished capacity to bind HLA class I. Correlating with their balance in human populations, B haplotypes favour reproductive success, whereas A haplotypes favour successful immune defence. Evolution of the B KIR haplotypes is thus unique to the human species.

    View details for DOI 10.1098/rstb.2011.0266

    View details for Web of Science ID 000300390100005

    View details for PubMedID 22312047

  • Review: Immunogenetics of human placentation PLACENTA Parham, P., Norman, P. J., Abi-Rached, L., HILTON, H. G., Guethlein, L. A. 2012; 33: S71-S80


    Natural killer (NK) cells are a population of lymphocytes that function in both immune defense and reproduction. Diversifying NK cell phenotype and function are interactions between NK cell receptors and major histocompatibility complex (MHC) class I ligands. As a consequence of strong and variable selection these ligand-receptor systems are polymorphic, rapidly evolving, and considerably species-specific. Counterparts to the human system of HLA class I ligands and killer cell immunoglobulin-like receptors (KIR) are present only in apes and Old World monkeys. HLA-C, the dominant ligand for human KIR and the only polymorphic HLA class I expressed by trophoblast, is further restricted to humans and great apes. Even then, the human system appears qualitatively different from that of chimpanzees, in that it has evolved a genetic balance between particular groups of receptors and ligands that favor reproductive success and other groups of receptors and ligands that have been correlated with disordered placentation. Human populations that have survived successive episodes of epidemic disease and population bottlenecks maintain a breadth of diversity for KIR and HLA class I, implying that loss of such diversity disfavors long-term survival of a human population.

    View details for DOI 10.1016/j.placenta.2011.11.020

    View details for Web of Science ID 000301868000014

    View details for PubMedID 22177321

  • The Shaping of Modern Human Immune Systems by Multiregional Admixture with Archaic Humans SCIENCE Abi-Rached, L., Jobin, M. J., Kulkarni, S., McWhinnie, A., Dalva, K., Gragert, L., Babrzadeh, F., Gharizadeh, B., Luo, M., Plummer, F. A., Kimani, J., Carrington, M., Middleton, D., Rajalingam, R., Beksac, M., Marsh, S. G., Maiers, M., Guethlein, L. A., Tavoularis, S., Little, A., Green, R. E., Norman, P. J., Parham, P. 2011; 334 (6052): 89-94


    Whole genome comparisons identified introgression from archaic to modern humans. Our analysis of highly polymorphic human leukocyte antigen (HLA) class I, vital immune system components subject to strong balancing selection, shows how modern humans acquired the HLA-B*73 allele in west Asia through admixture with archaic humans called Denisovans, a likely sister group to the Neandertals. Virtual genotyping of Denisovan and Neandertal genomes identified archaic HLA haplotypes carrying functionally distinctive alleles that have introgressed into modern Eurasian and Oceanian populations. These alleles, of which several encode unique or strong ligands for natural killer cell receptors, now represent more than half the HLA alleles of modern Eurasians and also appear to have been later introduced into Africans. Thus, adaptive introgression of archaic alleles has significantly shaped modern human immune systems.

    View details for DOI 10.1126/science.1209202

    View details for Web of Science ID 000295580300046

    View details for PubMedID 21868630

    View details for PubMedCentralID PMC3677943

  • Variable NK Cell Receptors Exemplified by Human KIR3DL1/S1 JOURNAL OF IMMUNOLOGY Parham, P., Norman, P. J., Abi-Rached, L., Guethlein, L. A. 2011; 187 (1): 11-19


    Variegated expression of variable NK cell receptors for polymorphic MHC class I broadens the range of an individual's NK cell response and the capacity for populations and species to survive disease epidemics and population bottlenecks. On evolutionary time scales, this component of immunity is exceptionally dynamic, unstable, and short-lived, being dependent on coevolution of ligands and receptors subject to varying, competing selection pressures. Consequently these systems of variable NK cell receptors are largely species specific and have recruited different classes of glycoprotein, even within the primate order of mammals. Such disparity helps to explain substantial differences in NK cell biology between humans and animal models, for which the population genetics is largely ignored. KIR3DL1/S1, which recognizes the Bw4 epitope of HLA-A and -B and is the most extensively studied of the variable NK cell receptors, exemplifies how variation in all possible parameters of function is recruited to diversify the human NK cell response.

    View details for DOI 10.4049/jimmunol.0902332

    View details for Web of Science ID 000291799300005

    View details for PubMedID 21690332

  • Hunter-gatherer genomic diversity suggests a southern African origin for modern humans PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Henn, B. M., Gignoux, C. R., Jobin, M., Granka, J. M., Macpherson, J. M., Kidd, J. M., Rodriguez-Botigue, L., Ramachandran, S., Hon, L., Brisbin, A., Lin, A. A., Underhill, P. A., Comas, D., Kidd, K. K., Norman, P. J., Parham, P., Bustamante, C. D., Mountain, J. L., Feldman, M. W. 2011; 108 (13): 5154-5162


    Africa is inferred to be the continent of origin for all modern human populations, but the details of human prehistory and evolution in Africa remain largely obscure owing to the complex histories of hundreds of distinct populations. We present data for more than 580,000 SNPs for several hunter-gatherer populations: the Hadza and Sandawe of Tanzania, and the ≠Khomani Bushmen of South Africa, including speakers of the nearly extinct N|u language. We find that African hunter-gatherer populations today remain highly differentiated, encompassing major components of variation that are not found in other African populations. Hunter-gatherer populations also tend to have the lowest levels of genome-wide linkage disequilibrium among 27 African populations. We analyzed geographic patterns of linkage disequilibrium and population differentiation, as measured by F(ST), in Africa. The observed patterns are consistent with an origin of modern humans in southern Africa rather than eastern Africa, as is generally assumed. Additionally, genetic variation in African hunter-gatherer populations has been significantly affected by interaction with farmers and herders over the past 5,000 y, through both severe population bottlenecks and sex-biased migration. However, African hunter-gatherer populations continue to maintain the highest levels of genetic diversity in the world.

    View details for DOI 10.1073/pnas.1017511108

    View details for Web of Science ID 000288894800009

    View details for PubMedID 21383195

  • Although Divergent in Residues of the Peptide Binding Site, Conserved Chimpanzee Patr-AL and Polymorphic Human HLA-A*02 Have Overlapping Peptide-Binding Repertoires JOURNAL OF IMMUNOLOGY Gleimer, M., Wahl, A. R., Hickman, H. D., Abi-Rached, L., Norman, P. J., Guethlein, L. A., Hammond, J. A., Draghi, M., Adams, E. J., Juo, S., Jalili, R., Gharizadeh, B., Ronaghi, M., Garcia, K. C., Hildebrand, W. H., Parham, P. 2011; 186 (3): 1575-1588


    Patr-AL is an expressed, non-polymorphic MHC class I gene carried by ∼50% of chimpanzee MHC haplotypes. Comparing Patr-AL(+) and Patr-AL(-) haplotypes showed Patr-AL defines a unique 125-kb genomic block flanked by blocks containing classical Patr-A and pseudogene Patr-H. Orthologous to Patr-AL are polymorphic orangutan Popy-A and the 5' part of human pseudogene HLA-Y, carried by ∼10% of HLA haplotypes. Thus, the AL gene alternatively evolved in these closely related species to become classical, nonclassical, and nonfunctional. Although differing by 30 aa substitutions in the peptide-binding α(1) and α(2) domains, Patr-AL and HLA-A*0201 bind overlapping repertoires of peptides; the overlap being comparable with that between the A*0201 and A*0207 subtypes differing by one substitution. Patr-AL thus has the A02 supertypic peptide-binding specificity. Patr-AL and HLA-A*0201 have similar three-dimensional structures, binding peptides in similar conformation. Although comparable in size and shape, the B and F specificity pockets of Patr-AL and HLA-A*0201 differ in both their constituent residues and contacts with peptide anchors. Uniquely shared by Patr-AL, HLA-A*0201, and other members of the A02 supertype are the absence of serine at position 9 in the B pocket and the presence of tyrosine at position 116 in the F pocket. Distinguishing Patr-AL from HLA-A*02 is an unusually electropositive upper face on the α(2) helix. Stimulating PBMCs from Patr-AL(-) chimpanzees with B cells expressing Patr-AL produced potent alloreactive CD8 T cells with specificity for Patr-AL and no cross-reactivity toward other MHC class I molecules, including HLA-A*02. In contrast, PBMCs from Patr-AL(+) chimpanzees are tolerant of Patr-AL.

    View details for DOI 10.4049/jimmunol.1002990

    View details for Web of Science ID 000286381200037

    View details for PubMedID 21209280

  • Different Patterns of Evolution in the Centromeric and Telomeric Regions of Group A and B Haplotypes of the Human Killer Cell Ig-Like Receptor Locus PLOS ONE Pyo, C., Guethlein, L. A., Vu, Q., Wang, R., Abi-Rached, L., Norman, P. J., Marsh, S. G., Miller, J. S., Parham, P., Geraghty, D. E. 2010; 5 (12)


    The fast evolving human KIR gene family encodes variable lymphocyte receptors specific for polymorphic HLA class I determinants. Nucleotide sequences for 24 representative human KIR haplotypes were determined. With three previously defined haplotypes, this gave a set of 12 group A and 15 group B haplotypes for assessment of KIR variation. The seven gene-content haplotypes are all combinations of four centromeric and two telomeric motifs. 2DL5, 2DS5 and 2DS3 can be present in centromeric and telomeric locations. With one exception, haplotypes having identical gene content differed in their combinations of KIR alleles. Sequence diversity varied between haplotype groups and between centromeric and telomeric halves of the KIR locus. The most variable A haplotype genes are in the telomeric half, whereas the most variable genes characterizing B haplotypes are in the centromeric half. Of the highly polymorphic genes, only the 3DL3 framework gene exhibits a similar diversity when carried by A and B haplotypes. Phylogenetic analysis and divergence time estimates, point to the centromeric gene-content motifs that distinguish A and B haplotypes having emerged ~6 million years ago, contemporaneously with the separation of human and chimpanzee ancestors. In contrast, the telomeric motifs that distinguish A and B haplotypes emerged more recently, ~1.7 million years ago, before the emergence of Homo sapiens. Thus the centromeric and telomeric motifs that typify A and B haplotypes have likely been present throughout human evolution. The results suggest the common ancestor of A and B haplotypes combined a B-like centromeric region with an A-like telomeric region.

    View details for DOI 10.1371/journal.pone.0015115

    View details for Web of Science ID 000285793200015

    View details for PubMedID 21206914

  • Primate-specific regulation of natural killer cells 27th Annual Symposium on Non-Human Primate Models for AIDS Parham, P., Abi-Rached, L., Matevosyan, L., Moesta, A. K., Norman, P. J., Aguilar, A. M., Guethlein, L. A. WILEY-BLACKWELL PUBLISHING, INC. 2010: 194–212


    Natural killer (NK) cells are circulating lymphocytes that function in innate immunity and placental reproduction. Regulating both development and function of NK cells is an array of variable and conserved receptors that interact with major histocompatibility complex (MHC) class I molecules. Families of lectin-like and immunoglobulin-like receptors are determined by genes in the natural killer complex (NKC) and leukocyte receptor complex (LRC), respectively. As a consequence of the strong, varying pressures on the immune and reproductive systems, NK cell receptors and their MHC class I ligands evolve rapidly, are highly diverse and exhibit dramatic species-specific differences. The variable, polymorphic family of killer cell immunoglobulin-like receptors (KIR) that regulate human NK cell development and function arose recently, from a single-copy gene during the evolution of simian primates. Our studies of KIR and MHC class I genes in representative species show how these two unlinked but functionally intertwined genetic complexes have co-evolved. In humans, combinations of KIR and HLA class I factors are associated with infectious diseases, including HIV/AIDS, autoimmunity, reproductive success and the outcome of therapeutic transplantation. The extraordinary, and unanticipated, divergence of human NK cell receptors and MHC class I ligands from their mouse counterparts can in part explain the difficulties experienced in finding informative mouse models for human diseases. Non-human primate models have far greater potential, but to realize their promise will first require more complete definition of the genetics and function of KIR and MHC variation in non-human primate species, at a level comparable to that achieved for the human species.

    View details for DOI 10.1111/j.1600-0684.2010.00432.x

    View details for Web of Science ID 000279448900003

    View details for PubMedID 20618586

  • Co-evolution of KIR2DL3 with HLA-C in a human population retaining minimal essential diversity of KIR and HLA class I ligands PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Gendzekhadze, K., Norman, P. J., Abi-Rached, L., Graef, T., Moesta, A. K., Layrisse, Z., Parham, P. 2009; 106 (44): 18692-18697


    Natural killer (NK) cells contribute to immunity and reproduction. Guiding these functions, and NK cell education, are killer cell Ig-like receptors (KIR), NK cell receptors that recognize HLA class I. In most human populations, these highly polymorphic receptors and ligands combine with extraordinary diversity. To assess how much of this diversity is necessary, we studied KIR and HLA class I at high resolution in the Yucpa, a small South Amerindian population that survived an approximate 15,000-year history of population bottleneck and epidemic infection, including recent viral hepatitis. The Yucpa retain the three major HLA epitopes recognized by KIR. Through balancing selection on a few divergent haplotypes the Yucpa maintain much of the KIR variation found worldwide. HLA-C*07, the strongest educator of C1-specific NK cells, has reached unusually high frequency in the Yucpa. Concomitantly, weaker variants of the C1 receptor, KIR2DL3, were selected and have largely replaced the form of KIR2DL3 brought by the original migrants from Asia. HLA-C1 and KIR2DL3 homozygosity has previously been correlated with resistance to viral hepatitis. Selection of weaker forms of KIR2DL3 in the Yucpa can be seen as compensation for the high frequency of the potent HLA-C*07 ligand. This study provides an estimate of the minimal KIR-HLA system essential for long-term survival of a human population. That it contains all functional elements of KIR diversity worldwide, attests to the competitive advantage it provides, not only for surviving epidemic infections, but also for rebuilding populations once infection has passed.

    View details for DOI 10.1073/pnas.0906051106

    View details for Web of Science ID 000271429800051

    View details for PubMedID 19837691

  • KIR2DS4 is a product of gene conversion with KIR3DL2 that introduced specificity for HLA-A*11 while diminishing avidity for HLA-C JOURNAL OF EXPERIMENTAL MEDICINE Graef, T., Moesta, A. K., Norman, P. J., Abi-Rached, L., Vago, L., Aguilar, A. M., Gleimer, M., Hammond, J. A., Guethlein, L. A., Bushnell, D. A., Robinson, P. J., Parham, P. 2009; 206 (11): 2557-2572


    Human killer cell immunoglobulin-like receptors (KIRs) are distinguished by expansion of activating KIR2DS, whose ligands and functions remain poorly understood. The oldest, most prevalent KIR2DS is KIR2DS4, which is represented by a variable balance between "full-length" and "deleted" forms. We find that full-length 2DS4 is a human histocompatibility leukocyte antigen (HLA) class I receptor that binds specifically to subsets of C1+ and C2+ HLA-C and to HLA-A*11, whereas deleted 2DS4 is nonfunctional. Activation of 2DS4+ NKL cells was achieved with A*1102 as ligand, which differs from A*1101 by unique substitution of lysine 19 for glutamate, but not with A*1101 or HLA-C. Distinguishing KIR2DS4 from other KIR2DS is the proline-valine motif at positions 71-72, which is shared with KIR3DL2 and was introduced by gene conversion before separation of the human and chimpanzee lineages. Site-directed swap mutagenesis shows that these two residues are largely responsible for the unique HLA class I specificity of KIR2DS4. Determination of the crystallographic structure of KIR2DS4 shows two major differences from KIR2DL: displacement of contact loop L2 and altered bonding potential because of the substitutions at positions 71 and 72. Correlation between the worldwide distributions of functional KIR2DS4 and HLA-A*11 points to the physiological importance of their mutual interaction.

    View details for DOI 10.1084/jem.20091010

    View details for Web of Science ID 000271164000022

    View details for PubMedID 19858347

  • Dimorphic Motifs in D0 and D1+D2 Domains of Killer Cell Ig-Like Receptor 3DL1 Combine to Form Receptors with High, Moderate, and No Avidity for the Complex of a Peptide Derived from HIV and HLA-A*2402 JOURNAL OF IMMUNOLOGY Sharma, D., Bastard, K., Guethlein, L. A., Norman, P. J., Yawata, N., Yawata, M., Pando, M., Thananchai, H., Dong, T., Rowland-Jones, S., Brodsky, F. M., Parham, P. 2009; 183 (7): 4569-4582


    Comparison of mutant killer cell Ig-like receptor (KIR) 3DL1*015 substituted at natural positions of variation showed that tryptophan/leucine dimorphism at position 283 uniquely changes receptor conformation and can strongly influence binding of the A24nef tetramer. Dimorphic motifs at positions 2, 47, and 54 in D0 and 182 and 283 in D1+D2 distinguish the two 3DL1 lineages, typified by 3DL1*005 and 3DL1*015. The interlineage recombinant, KIR3DL1*001, combines D0 of 3DL1*005 with D1+D2 of 3DL1*015 and binds A24nef more strongly than either parent. In contrast, the reciprocal recombinant with D0 from 3DL1*015 and D1+D2 from 3DL1*005 cannot bind A24nef. Thus, D0 polymorphism directly affects the avidity of the KIR3DL1 ligand binding site. From these observations, multiple sequence alignment, and homology modeling, we constructed structural models for KIR3DL1 and its complex with A24nef. In these models, D0, D1, and D2 come together to form a binding surface for A24nef, which is contacted by all three Ig-like domains. A central pocket binds arginine 83, the only Bw4 motif residue essential for KIR3DL1 interaction, similar to the binding of lysine 80 in HLA-C by KIR2DL1. Central to this interaction is a salt bridge between arginine 83 of Bw4 and glutamate 282 of 3DL1, which juxtaposes the functionally influential dimorphism at position 283. Further 3DL1 mutants were tested and shown to have A24nef-binding properties consistent with the models. A24nef was not bound by KIR3DS1, the activating counterpart of KIR3DL1. Moreover, introducing any one of three residues specific to KIR3DS1, serine 163, arginine 166, or leucine 199, into 3DL1*015, abrogated A24nef binding.

    View details for DOI 10.4049/jimmunol.0901734

    View details for Web of Science ID 000270522500052

    View details for PubMedID 19752231

  • Killer Ig-Like Receptor (KIR) Genotype Predicts the Capacity of Human KIR-Positive CD56(dim) NK Cells to Respond to Pathogen-Associated Signals JOURNAL OF IMMUNOLOGY Korbel, D. S., Norman, P. J., Newman, K. C., Horowitz, A., Gendzekhadze, K., Parham, P., Riley, E. M. 2009; 182 (10): 6426-6434


    IFN-gamma emanating from NK cells is an important component of innate defense against infection. In this study, we demonstrate that, following in vitro stimulation of human peripheral blood NK cells with a variety of microbial ligands, CD56(dim) as well as CD56(bright) NK cells contribute to the overall NK cell IFN-gamma response with, for most cell donors, IFN-gamma(+) CD56(dim) NK cells outnumbering IFN-gamma(+) CD56(bright) NK cells. We also observe that the magnitude of the human NK IFN-gamma response to microbial ligands varies between individuals; that the antimicrobial response of CD56(bright), but not CD56(dim), NK cells is highly correlated with that of myeloid accessory cells; and that the ratio of IFN-gamma(+) CD56(dim) to IFN-gamma(+) CD56(bright) NK cells following microbial stimulation differs between individuals but remains constant for a given donor over time. Furthermore, ratios of IFN-gamma(+) CD56(dim) to IFN-gamma(+) CD56(bright) NK cells for different microbial stimuli are highly correlated and the relative response of CD56(dim) and CD56(bright) NK cells is highly significantly associated with killer Ig-like receptor (KIR) genotype. These data reveal an influence of KIR genotype, possibly mediated via NK cell education, on the ability of NK cells to respond to nonviral infections and have implications for genetic regulation of susceptibility to infection in humans.

    View details for DOI 10.4049/jimmunol.0804224

    View details for Web of Science ID 000265899800061

    View details for PubMedID 19414796

  • Meiotic recombination generates rich diversity in NK cell receptor genes, alleles, and haplotypes GENOME RESEARCH Norman, P. J., Abi-Rached, L., Gendzekhadze, K., Hammond, J. A., Moesta, A. K., Sharma, D., Graef, T., McQueen, K. L., Guethlein, L. A., Carrington, C. V., Chandanayingyong, D., Chang, Y., Crespi, C., Saruhan-Direskeneli, G., Hameed, K., Kamkamidze, G., Koram, K. A., Layrisse, Z., Matamoros, N., Mila, J., Park, M. H., Pitchappan, R. M., Ramdath, D. D., Shiau, M., Stephens, H. A., Struik, S., Tyan, D., Verity, D. H., Vaughan, R. W., Davis, R. W., Fraser, P. A., Riley, E. M., Ronaghi, M., Parham, P. 2009; 19 (5): 757-769


    Natural killer (NK) cells contribute to the essential functions of innate immunity and reproduction. Various genes encode NK cell receptors that recognize the major histocompatibility complex (MHC) Class I molecules expressed by other cells. For primate NK cells, the killer-cell immunoglobulin-like receptors (KIR) are a variable and rapidly evolving family of MHC Class I receptors. Studied here is KIR3DL1/S1, which encodes receptors for highly polymorphic human HLA-A and -B and comprises three ancient allelic lineages that have been preserved by balancing selection throughout human evolution. While the 3DS1 lineage of activating receptors has been conserved, the two 3DL1 lineages of inhibitory receptors were diversified through inter-lineage recombination with each other and with 3DS1. Prominent targets for recombination were D0-domain polymorphisms, which modulate enhancer function, and dimorphism at position 283 in the D2 domain, which influences inhibitory function. In African populations, unequal crossing over between the 3DL1 and 3DL2 genes produced a deleted KIR haplotype in which the telomeric "half" was reduced to a single fusion gene with functional properties distinct from its 3DL1 and 3DL2 parents. Conversely, in Eurasian populations, duplication of the KIR3DL1/S1 locus by unequal crossing over has enabled individuals to carry and express alleles of all three KIR3DL1/S1 lineages. These results demonstrate how meiotic recombination combines with an ancient, preserved diversity to create new KIR phenotypes upon which natural selection acts. A consequence of such recombination is to blur the distinction between alleles and loci in the rapidly evolving human KIR gene family.

    View details for DOI 10.1101/gr.085738.108

    View details for Web of Science ID 000265668800009

    View details for PubMedID 19411600

  • Chimpanzees Use More Varied Receptors and Ligands Than Humans for Inhibitory Killer Cell Ig-Like Receptor Recognition of the MHC-C1 and MHC-C2 Epitopes JOURNAL OF IMMUNOLOGY Moesta, A. K., Abi-Rached, L., Norman, P. J., Parham, P. 2009; 182 (6): 3628-3637


    Humans and chimpanzees have orthologous MHC class I, but few orthologous killer cell Ig-like receptors (KIR). Most divergent are lineage III KIR, which in humans include the inhibitory KIR2DL1 and 2DL2/3 specific for HLA-C. Six lineage III chimpanzee KIR were identified as candidate inhibitory MHC-C receptors and studied using cytolytic assays, to assess the capacity of a defined KIR to function with a defined MHC class I allotype, and direct binding assays with KIR-Fc fusion proteins. Pt-KIR2DL6 and 2DL8 were demonstrated to be inhibitory C1 receptors with a specificity and specificity-determining residue (lysine 44) like KIR2DL3. Analogously, Pt-KIR2DL7 is like KIR2DL1, an inhibitory C2 receptor having methionine 44. Pt-KIR3DL4 and 3DL5 are unusual lineage III KIR with D0 domains, which are also inhibitory C2 receptors with methionine 44. Removal of D0 from KIR3DL, or its addition to KIR2DL, had no effect on KIR function. Pt-KIR2DL9, a fourth inhibitory C2 receptor, has glutamate 44, a previously uncharacterized specificity-determining residue that is absent from human KIR. Reconstruction of the ancestral hominoid KIR sequence shows it encoded lysine 44, indicating that KIR having methionine 44 and glutamate 44 subsequently evolved by independent point substitutions. Thus, MHC-C2-specific KIR have evolved independently on at least two occasions. None of the six chimpanzee KIR studied resembles KIR2DL2, which interacts strongly with C1 and cross-reacts with C2. Whereas human HLA-B allotypes that have functional C1 epitopes are either rare (HLA-B*73) or geographically localized (HLA-B*46), some 25% of Patr-B allotypes have the C1 epitope and are functional KIR ligands.

    View details for DOI 10.4049/jimmunol.0803401

    View details for Web of Science ID 000264084100033

    View details for PubMedID 19265141

  • Polymorphic Sites Away from the Bw4 Epitope That Affect Interaction of Bw4(+) HLA-B with KIR3DL1 JOURNAL OF IMMUNOLOGY Sanjanwala, B., Draghi, M., Norman, P. J., Guethlein, L. A., Parham, P. 2008; 181 (9): 6293-6300


    KIR3DL1 is a polymorphic, inhibitory NK cell receptor specific for the Bw4 epitope carried by subsets of HLA-A and HLA-B allotypes. The Bw4 epitope of HLA-B*5101 and HLA-B*1513 is determined by the NIALR sequence motif at positions 77, 80, 81, 82, and 83 in the alpha(1) helix. Mutation of these positions to the residues present in the alternative and nonfunctional Bw6 motif showed that the functional activity of the Bw4 epitopes of B*5101 and B*1513 is retained after substitution at positions 77, 80, and 81, but lost after substitution of position 83. Mutation of leucine to arginine at position 82 led to loss of function for B*5101 but not for B*1513. Further mutagenesis, in which B*1513 residues were replaced by their B*5101 counterparts, showed that polymorphisms in all three extracellular domains contribute to this functional difference. Prominent were positions 67 in the alpha(1) domain, 116 in the alpha(2) domain, and 194 in the alpha(3) domain. Lesser contributions were made by additional positions in the alpha(2) domain. These positions are not part of the Bw4 epitope and include residues shaping the B and F pockets that determine the sequence and conformation of the peptides bound by HLA class I molecules. This analysis shows how polymorphism at sites throughout the HLA class I molecule can influence the interaction of the Bw4 epitope with KIR3DL1. This influence is likely mediated by changes in the peptides bound, which alter the conformation of the Bw4 epitope.

    View details for Web of Science ID 000260659000055

    View details for PubMedID 18941220

  • Novel KIR3DL1 alleles and their expression levels on NK cells: Convergent evolution of KIR3DL1 phenotype variation? JOURNAL OF IMMUNOLOGY Thomas, R., Yamada, E., Alter, G., Martin, M. P., Bashirova, A. A., Norman, P. J., Altfeld, M., Parham, P., Anderson, S. K., McVicar, D. W., Carrington, M. 2008; 180 (10): 6743-6750


    KIR3DL1 shows extensive polymorphism, and its variation has functional significance in terms of cell-surface expression levels and inhibitory capacity. We characterized nine KIR3DL1 alleles (*022, *028, *029, *033, *035, *051, *052, *053, and *054), four of which were identified for the first time in this study, and compared them to known alleles in phylogenetic analysis. Blood was available from eight individuals with these alleles, and cell-surface expression on NK cells could be determined for six of them using the KIR3DL1-specific Ab DX9. Four of the alleles were expressed at clearly detectable levels, and two others showed exceptionally low levels of expression. Site-directed mutagenesis demonstrated that single amino acid changes can result in either diminished or enhanced DX9 staining compared with the respective related KIR3DL1 allotypes. These results raise the possibility that KIR3DL1 evolution maintains variation in KIR3DL1 cell-surface expression levels, potentially due to the effect of such variation on functional capacity.

    View details for Web of Science ID 000257507100039

    View details for PubMedID 18453594

  • Synergistic polymorphism at two positions distal to the ligand-binding site makes KIR2DL2 a stronger receptor for HLA-C than KIR2DL3 JOURNAL OF IMMUNOLOGY Moesta, A. K., Norman, P. J., Yawata, M., Yawata, N., Gleimer, M., Parham, P. 2008; 180 (6): 3969-3979


    Interactions between HLA-C ligands and inhibitory killer cell Ig-like receptors (KIR) control the development and response of human NK cells. This regulatory mechanism is usually described by mutually exclusive interactions of KIR2DL1 with C2 having lysine 80, and KIR2DL2/3 with C1 having asparagine 80. Consistent with this simple rule, we found from functional analysis and binding assays to 93 HLA-A, HLA-B, and HLA-C isoforms that KIR2DL1*003 bound all C2, and only C2, allotypes. The allotypically related KIR2DL2*001 and KIR2DL3*001 interacted with all C1, but they violated the simple rule through interactions with several C2 allotypes, notably Cw*0501 and Cw*0202, and two HLA-B allotypes (B*4601 and B*7301) that share polymorphisms with HLA-C. Although the specificities of the "cross-reactions" were similar for KIR2DL2*001 and KIR2DL3*001, they were stronger for KIR2DL2*001, as were the reactions with C1. Mutagenesis explored the avidity difference between KIR2DL2*001 and KIR2DL3*001. Recombinant mutants mapped the difference to the Ig-like domains, where site-directed mutagenesis showed that the combination, but not the individual substitutions, of arginine for proline 16 in D1 and cysteine for arginine 148 in D2 made KIR2DL2*001 a stronger receptor than KIR2DL3*001. Neither residue 16 or 148 is part of, or near to, the ligand-binding site. Instead, their juxtaposition near the flexible hinge between D1 and D2 suggests that their polymorphisms affect the ligand-binding site by changing the hinge angle and the relative orientation of the two domains. This study demonstrates how allelic polymorphism at sites distal to the ligand-binding site of KIR2DL2/3 has diversified this receptor's interactions with HLA-C.

    View details for Web of Science ID 000257506600044

    View details for PubMedID 18322206

  • Unusual selection on the KIR3DL1/S1 natural killer cell receptor in Africans NATURE GENETICS Norman, P. J., Abi-Rached, L., Gendzekhadze, K., Korbel, D., Gleimer, M., Rowley, D., Bruno, D., Carrington, C. V., Chandanayingyong, D., Chang, Y., Crespi, C., Saruhan-Direskeneli, G., Fraser, P. A., Hameed, K., Kamkamidze, G., Koram, K. A., Layrisse, Z., Matamoros, N., Mila, J., Park, M. H., Pitchappan, R. M., Ramdath, D. D., Shiau, M., Stephens, H. A., Struik, S., Verity, D. H., Vaughan, R. W., Tyan, D., Davis, R. W., Riley, E. M., Ronaghi, M., Parham, P. 2007; 39 (9): 1092-1099


    Interactions of killer cell immunoglobulin-like receptors (KIRs) with major histocompatibility complex (MHC) class I ligands diversify natural killer cell responses to infection. By analyzing sequence variation in diverse human populations, we show that the KIR3DL1/S1 locus encodes two lineages of polymorphic inhibitory KIR3DL1 allotypes that recognize Bw4 epitopes of protein">HLA-A and HLA-B and one lineage of conserved activating KIR3DS1 allotypes, also implicated in Bw4 recognition. Balancing selection has maintained these three lineages for over 3 million years. Variation was selected at D1 and D2 domain residues that contact HLA class I and at two sites on D0, the domain that enhances the binding of KIR3D to HLA class I. HLA-B variants that gained Bw4 through interallelic microconversion are also products of selection. A worldwide comparison uncovers unusual KIR3DL1/S1 evolution in modern sub-Saharan Africans. Balancing selection is weak and confined to D0, KIR3DS1 is rare and KIR3DL1 allotypes with similar binding sites predominate. Natural killer cells express the dominant KIR3DL1 at a high frequency and with high surface density, providing strong responses to cells perturbed in Bw4 expression.

    View details for DOI 10.1038/ng2111

    View details for Web of Science ID 000249122400018

    View details for PubMedID 17694054

  • Episodes of natural selection shaped the interactions of IgA-Fc with Fc alpha RI and bacterial decoy proteins JOURNAL OF IMMUNOLOGY Abi-Rached, L., Dorighi, K., Norman, P. J., Yawata, M., Parham, P. 2007; 178 (12): 7943-7954


    FcalphaRI, a receptor for IgA-Fc, recruits myeloid cells to attack IgA-coated pathogens. By competing with FcalphaRI for IgA, bacterial decoys, like SSL7 of Staphylococcus aureus, subvert this defense. We examined how pathogen selection has driven the diversification and coevolution of IgA and FcalphaRI. In higher primates, the IgA binding site of FcalphaRI diversified under positive selection, a strong episode occurring in hominoid ancestors about the time of the IgA gene duplication. The differential binding of SSL7 to IgA-Fc of different species correlates with substitution at seven positions in IgA-Fc, two of which were positively selected in higher primates. Two others, which reduce SSL7 binding, emerged during episodes of positive selection in the rabbit and rodent lineages. The FcalphaRI-IgA interaction evolves episodically under two types of positive selection: pressure from pathogen decoys selects for IgA escape variants which, in turn, selects for FcalphaRI variants to keep up with the novel IgA. When FcalphaRI cannot keep up, its function is lost and the gene becomes susceptible to elimination, as occurred in the mouse genome, either by chance or selection on one of the many linked, variable immune system genes. A cluster of positively selected residues presents a putative binding site for unknown IgA-binding factors.

    View details for Web of Science ID 000247189500053

    View details for PubMedID 17548632

  • Reduced telomere length in rheumatoid arthritis is independent of disease activity and duration ANNALS OF THE RHEUMATIC DISEASES Steer, S. E., Williams, F. M., Kato, B., Gardner, J. P., Norman, P. J., Hall, M. A., Kimura, M., Vaughan, R., Aviv, A., Spector, T. D. 2007; 66 (4): 476-480


    Rheumatoid arthritis (RA) is associated with reduced lifespan and shortened telomere length in lymphocytes, but the mechanism underlying this is unclear. Telomere loss in white blood cells (WBC) is accelerated by oxidative stress and inflammation in vitro. It was postulated that the accelerated WBC telomere shortening in RA occurs as a result of exposure to chronic inflammation.To measure telomere terminal restriction fragment (TRF) length in a large cohort of RA cases and healthy controls, to explore associations of TRF length with features of disease and with RA-associated HLA-DRB1 alleles.WBC and TRF length were measured by Southern blot in DNA from 176 hospital-based RA cases satisfying the 1987 American College of Rheumatology criteria and from 1151 controls. TRF length was compared between cases and controls, and the effects of disease duration, severity and HLA-DRB1 alleles encoding the shared epitope (SE) were assessed.Age- and sex-adjusted TRF length was significantly shorter in RA cases compared with controls (p<0.001). There was no association between age- and sex-adjusted TRF length and disease duration, C reactive protein or Larsen score. The presence of one or more SE-encoding alleles was associated with reduced adjusted TRF length in RA cases (SE positive vs SE negative cases, p=0.038), but not in controls.The reduced TRF length in a large group of patients with RA compared with controls has been shown. The reduction is apparently independent of disease duration and markers of disease severity, but is influenced by HLA-DRB1 genotype.

    View details for DOI 10.1136/ard.2006.059188

    View details for Web of Science ID 000244924700009

    View details for PubMedID 17114192

  • High KIR diversity in Amerindians is maintained using few gene-content haplotypes IMMUNOGENETICS Gendzekhadze, K., Norman, P. J., Abi-Rached, L., Layrisse, Z., Parham, P. 2006; 58 (5-6): 474-480


    Interaction between killer cell immunoglobulin-like receptors (KIR) and cognate HLA class I ligands influences the innate and adaptive immune response to infection. The KIR family varies in gene content and allelic polymorphism, thereby, distinguishing individuals and populations. KIR gene content was determined for 230 individuals from three Amerindian tribes from Venezuela: the Yucpa, Bari and Warao. Gene-content haplotypes could be assigned to 212 individuals (92%) because only five different haplotypes were present-group A and four group B. Six different haplotype combinations accounted for >80% of individuals. Each tribe has distinctive genotype frequencies. Despite few haplotypes, all 14 KIR genes are at high frequency in the three tribes, with the exception of 2DS3. Each population has an even frequency of group A and B haplotypes. Allele-level analysis of 3DL1/S1 distinguished five group A haplotypes and six group B haplotypes. The high frequency and divergence of the KIR haplotypes in the Amerindian tribes provide greater KIR diversity than is present in many larger populations. An extreme case being the Yucpa, for whom two gene-content haplotypes account for >90% of the population. These comprise the group A haplotype and a group B haplotype containing all the KIR genes, except 2DS3, that typify the group B haplotypes. Here is clear evidence for balancing selection on the KIR system and the biological importance of both A and B haplotypes for the survival of human populations.

    View details for DOI 10.1007/s00251-006-0108-3

    View details for Web of Science ID 000238014900017

    View details for PubMedID 16738943

  • Complex interactions: The immunogenetics of human leukocyte antigen and killer cell immunoglobulin-like receptors SEMINARS IN HEMATOLOGY Norman, P. J., Parham, P. 2005; 42 (2): 65-75


    The killer cell immunoglobulin-like receptors (KIR) for human leukocyte antigen (HLA) modulate innate and adaptive immunity by controlling effector cells. HLA and KIR are encoded in genomic regions that have complex organization and exhibit exceptional diversity within and among human population groups. This diversity is likely to have arisen to combat a constantly evolving pathogen challenge. Numerous variations influence the expression level or function of KIR molecules and can affect their interaction with HLA, with important implications for the immune response. The functional variety of natural immune responses that are controlled by HLA and KIR interactions is genetically determined and maintained by natural selection.

    View details for DOI 10.1053/j.seminhematol.2005.01.007

    View details for Web of Science ID 000228661400002

    View details for PubMedID 15846572

  • Isolation, purification and flow cytometric analysis of human intrahepatic lymphocytes using an improved technique LABORATORY INVESTIGATION Morsy, M. A., Norman, P. J., Mitry, R., Rela, M., Heaton, N. D., Vaughan, R. W. 2005; 85 (2): 285-296


    Intrahepatic lymphocytes (IHL) with their diverse and distinctive subsets emphasise the importance of the liver as a site of immunological activity, but special care is required for their isolation and characterisation. Protocols for IHL isolation, purification and FACS analysis were devised and compared with published extraction protocols. We have reduced the time that IHL are exposed to potentially damaging enzymes during extraction and purified specific subsets using monoclonal antibody (mAb)-coated magnetic microbeads. This has yielded IHL populations with higher viability than previously described protocols (92-95%, compared with 39-86%). Flow cytometric characterisation of IHL subset immunophenotypes was optimised by combining CD45 staining (fluorescence gating) with traditional light scatter properties. Using a panel of mAb and liver biopsies obtained from 23 cadaveric liver transplant donors, we show that the normal liver contains a heterogeneous IHL population with distinctive phenotypes. CD8+ IHL was the predominant population with a mean CD4/CD8 ratio of 1:1.7. Up to 40% of IHL expressed gammadeltaTCR and a third expressed CD56 NK marker; indicating a site of intense immunological activity. The techniques described will allow these cell types to be isolated, fully characterised and their physiological functions to be determined. The histologically normal liver contains heterogeneous and diverse IHL with large numbers of CD8+, NK, NKT and gammadelta+ cells.

    View details for DOI 10.1038/labinvest.3700219

    View details for Web of Science ID 000226719900015

    View details for PubMedID 15640833

  • KIR gene Allele frequencies in populations from USA (African American) USA (European) and England HUMAN IMMUNOLOGY Carrington, M., Gao, X., Norman, P. 2004; 65 (9-10): 1191-1191
  • SNP haplotypes and allele frequencies show evidence for disruptive and balancing selection in the human leukocyte receptor complex IMMUNOGENETICS Norman, P. J., Cook, M. A., Carey, B. S., Carrington, C. V., Verity, D. H., Hameed, K., Ramdath, D. D., Chandanayingyong, D., Leppert, M., Stephens, H. A., Vaughan, R. W. 2004; 56 (4): 225-237


    The human leukocyte receptor complex (LRC) of Chromosome 19q13.4 encodes polymorphic and highly homologous genes that are expressed by cells of the immune system and regulate their function. There is an enormous diversity at the LRC, most particularly the variable number of killer cell immunoglobulin-like receptor (KIR) genes. KIR have been associated with several disease processes due to their interaction with polymorphic human leukocyte antigen class I molecules. We have assessed haplotype compositions, linkage disequilibrium patterns and allele frequencies in two Caucasoid population samples (n=54, n=100), using a composite of single-nucleotide polymorphism (SNP) markers and high-resolution, allele-specific molecular genotyping. Particular KIR loci segregated with SNP and other markers, forming two blocks that were separated by a region with a greater history of recombination. The KIR haplotype composition and allele frequency distributions were consistent with KIR having been subject to balancing selection (Watterson's F: P=0.001). In contrast, there was a high inter-population heterogeneity measure for the LRC-encoded leukocyte immunoglobulin-like receptor A3 (LILRA3), indicating pathogen-driven disruptive selection (Wright's FST=0.32). An assessment of seven populations representative of African, Asian and Caucasoid ethnic groups (total n=593) provided little evidence for long-range LRC haplotypes. The different natural selection pressures acting on each locus may have contributed to a lack of linkage disequilibrium between them.

    View details for DOI 10.1007/s00251-004-0674-1

    View details for Web of Science ID 000223124200001

    View details for PubMedID 15185041

  • Natural killer-cell activity after human renal transplantation in relation to killer immunoglobulin-like receptors and human leukocyte antigen mismatch TRANSPLANTATION Vampa, M. L., Norman, P. J., Burnapp, L., Vaughan, R. W., Sacks, S. H., Wong, W. 2003; 76 (8): 1220-1228


    Natural killer (NK) cells use killer immunoglobulin-like receptors (KIR) that bind to self-class I major histocompatibility complex (MHC) molecules to prevent killing of autologous cells. Mismatched allografts, which do not express recipient MHC class I molecules, can therefore be potential targets for NK-cell killing. In our living related-unrelated renal transplantation program, donor-recipient pairs vary in the amount of both HLA and KIR genes they share. This provides us with a unique opportunity to dissect the influence of KIR on NK-cell function after transplantation.Recipient NK cells were used in a cytotoxicity assay against donor peripheral blood mononuclear cells 2 days before, on the day of, and 3 days after transplantation. Results were correlated to HLA-KIR compatibility between donor and recipient.NK killing, in a direct ex vivo setting, was demonstrated to be HLA mismatch dependent. Recipient NK antidonor cytotoxicity was unaltered despite having received 2 days' treatment with cyclosporine A before transplantation. However, cytotoxicity increased 3 days after transplantation in 71% of recipients. Recipients exhibiting increased NK cytotoxicity against their donors after transplantation were found to possess more activating KIR genes specific for donor class I MHC molecules than those in whom killing activity did not increase (P<0.04).NK cells are activated after transplantation despite quadruple immunosuppression, suggesting that recipient NK-cell cytotoxicity against the donor may be a previously unrecognized area of the rejection process, especially in poorly matched donor-recipient pairs where the recipient may not express the correct repertoire of inhibitory receptors to prevent killing of donor cells.

    View details for DOI 10.1097/01.TP.0000083896.91215.C7

    View details for Web of Science ID 000186269600015

    View details for PubMedID 14578757

  • Analysis of Fc gamma receptor II (CD32) polymorphism in populations of African and South Asian ancestry reveals east-west geographic gradients of allele frequencies EUROPEAN JOURNAL OF IMMUNOGENETICS Carrington, C. V., Norman, P. J., Vaughan, R. W., Kondeatis, E., Ramdath, D. D., Hameed, K., Stephens, H. A. 2003; 30 (5): 375-379


    Analysis of FcgammaRIIA alleles in Pakistanis and in Trinidadians of South Asian, African and mixed ancestry revealed no significant differences between Trinidadian South Asians and Pakistanis. H131 homozygotes were more common among Trinidadian South Asians than among Africans and those of mixed ancestry. Comparison with other populations revealed east-west geographic gradients of allele frequencies.

    View details for Web of Science ID 000186179500009

    View details for PubMedID 14641546

  • DNA sequence variation and molecular genotyping of natural killer leukocyte immunoglobulin-like receptor, LILRA3 IMMUNOGENETICS Norman, P. J., Carey, B. S., Stephens, H. A., Vaughan, R. W. 2003; 55 (3): 165-171


    Leukocyte immunoglobulin-like receptors (LILRs) resemble killer cell immunoglobulin-like receptors (KIR) in structure and function and the KIR and LILR gene families form the major part of the leukocyte receptor cluster (LRC) of human chromosome 19q13.4. Unlike KIR, the LILR gene clusters do not vary in gene number. However, some individuals lack expression of LILRA3. This null allele has a 6.7-kb deletion, which encompasses the first six translated exons. This haplotype enabled unambiguous direct sequencing of LILRA3 alleles using genomic DNA from individuals heterozygous for the deletion. We have performed nucleotide sequencing of a 2.5-kb region within LILRA3 and identified eight bi-allelic substitutions, four of which were non-synonymous. Two from four previously identified LILRA3 cDNA sequences were confirmed and a further six alleles characterised, of which four will encode unique peptides. At least one of the polymorphic positions identified (encoding residue 84 of the first Ig domain) is likely to directly influence ligand binding. A PCR-SSP molecular genotyping system was developed and used to describe a panel of 172 Caucasoid individuals from South-East England. Six alleles were present in this group but they were unevenly distributed, as three alleles accounted for 88% of the studied chromosomes.

    View details for DOI 10.1007/s00251-003-0561-1

    View details for Web of Science ID 000183636900005

    View details for PubMedID 12750859

  • A multi-laboratory characterization of the KIR genotypes of 10th International Histocompatibility Workshop cell lines HUMAN IMMUNOLOGY Cook, M. A., Norman, P. J., Curran, M. D., Maxwell, L. D., Briggs, D. C., Middleton, D., Vaughan, R. W. 2003; 64 (5): 567-571


    Killer immunoglobulinlike receptors (KIRs) are expressed on natural killer and T cells. Both inhibitory and noninhibitory forms have been described, leading to inhibition or continuation of cellular killing activity. The natural ligands identified so far of KIRs are class I human leukocyte antigens (HLA). In particular, the interaction of some KIRs with HLA-Cw has been well characterized. Recent work has implicated KIRs in affecting the outcome of hematopoietic stem-cell transplant (HSCT). This may well lead to a requirement for prospective KIR typing of donor and recipient. We have utilized different typing systems (two using polymerase chain reaction-sequence-specific primers, and one using polymerase chain reaction-sequence-specific oligonucleotide probes) in three separate laboratories to characterize the KIR gene complement of 25 cell lines from the 10th International Histocompatibility Workshop. There were consistent results in 22, and minor differences in 3. When compared with previous results for some of these cell lines, no further differences were found. The differences are due to typing of KIRs KIR2DL1 and KIR2DS5, and may be explained by technical differences or the inability to type new variants. Further improvements in typing may be required if population and clinical studies are to produce accurate results.

    View details for DOI 10.1016/S0198-8859(03)00042-9

    View details for Web of Science ID 000182424500011

    View details for PubMedID 12691708

  • A comparison of HLA-DR and -DQ allele and haplotype frequencies in trinidadian populations of African, South Asian, and mixed ancestry HUMAN IMMUNOLOGY Carrington, C. V., Kondeatis, E., Ramdath, D. D., Norman, P. J., Vaughan, R. W., Stephens, H. A. 2002; 63 (11): 1045-1054


    Using polymerase chain reaction-sequence-specific primer (PCR-SSP) typing, this study determined the frequency of human leukocyte antigen (HLA) DR- and -DQ alleles and haplotypes in individuals of African (n = 75), South Asian (n = 98), and mixed (n = 102) ancestry from the Caribbean island of Trinidad. We detected 19 different haplotypes containing DRB3, 8 containing DRB4, 6 containing DRB5, and 6 different haplotypes without DRB3/4/5 genes. Twenty-nine haplotypes were identified in Africans, 24 in the South Asians, and 32 in the mixed group. We detected significant differences between the populations, principally at the DQA1 and DQB1 loci, although the allele frequency for DRB1*0901 was highest in the Africans (p(c) < 0.05). Trinidad African and mixed groups were generally more diverse than the South Asians and displayed a wider range of DRB1-DQB1 associations; DQB1*02 and DQB1*0301 each associated with five to six different DRB1 alleles in the Africans and mixed group but only two in South Asians. In the Africans and the mixed group, DQB1*04 was found with DRB1*0302 and DRB1*04, but only with DRB1*08 in the South Asians. Trinidad Africans revealed consistencies with populations in Western, Central, and Northern Africa, but differed considerably from individual populations on the African continent. Trinidad South Asians displayed similar allele frequencies and associations to other populations from Northern India.

    View details for Web of Science ID 000178924200010

    View details for PubMedID 12392858

  • Analysis of candidate genes on chromosome 19 in coeliac disease: an association study of the KIR and LILR gene clusters EUROPEAN JOURNAL OF IMMUNOGENETICS Moodie, S. J., Norman, P. J., King, A. L., Fraser, J. S., Curtis, D., Ellis, H. J., Vaughan, R. W., Ciclitira, P. J. 2002; 29 (4): 287-291


    Coeliac disease is strongly heritable, with more than half of the genetic susceptibility estimated to come from genes outside the HLA region. Several candidate regions have been suggested from genome-wide linkage studies including chromosome 19q13.4 where linkage has been replicated between populations. The natural killer (NK) cell immunoglobulin-like receptors (KIRs) and leukocyte immunoglobulin-like receptor (LILR, also known as ILT and LIR) gene clusters lie within this region in the leukocyte receptor cluster (LRC). KIR molecules are involved in cytotoxic lymphocyte function and expressed by intraepithelial T and NK cells in the duodenum. We studied 132 unrelated UK Caucasian coeliac patients and their parents together with a control group of 171 UK Caucasians. PCR-SSP for KIR2DL1, KIR2DL2, KIR2DL3, KIR2DL5, LILRA3 (ILT6), LILRA3 deletion and an LILRA3 exon 3 single nucleotide polymorphism (SNP) allowed classification of KIR genotypes into five categories and determination of homozygosity or heterozygosity for the common A and B type KIR haplotypes (as defined in the text) and for the LILRA3 deletion. Case-control analysis found no association of the five KIR genotype categories, the A or B KIR haplotypes, the LILRA3 gene deletion or the LILRA3 exon 3 SNP with coeliac disease. A transmission disequilibrium test also found no association of the A and B KIR haplotypes or the LILRA3 gene deletion with coeliac disease.

    View details for Web of Science ID 000176797500002

    View details for PubMedID 12121272

  • Quantitative-trait loci on chromosomes 1, 2, 3, 4, 8, 9, 11, 12, and 18 control variation in levels of T and B lymphocyte subpopulations AMERICAN JOURNAL OF HUMAN GENETICS Hall, M. A., Norman, P. J., Thiel, B., Tiwari, H., Peiffer, A., Vaughan, R. W., Prescott, S., Leppert, M., Schork, N. J., Lanchbury, J. S. 2002; 70 (5): 1172-1182


    Lymphocyte subpopulation levels are used for prognosis and monitoring of a variety of human diseases, especially those with an infectious etiology. As a primary step to defining the major gene variation underlying these phenotypes, we conducted the first whole-genome screen for quantitative variation in lymphocyte count, CD4 T cell, CD8 T cell, B cell, and natural killer cell numbers, as well as CD4:CD8 ratio. The screen was performed in 15 of the CEPH families that form the main human genome genetic project mapping resource. Quantitative-trait loci (QTLs) that account for significant proportions of the phenotypic variance of lymphocyte subpopulations were detected on chromosomes 1, 2, 3, 4, 8, 9, 11, 12, and 18. The most significant QTL found was for CD4 levels on chromosome 8 (empirical P=.00005). Two regions of chromosome 4 showed significant linkage to CD4:CD8 ratio (empirical P=.00007 and P=.003). A QTL for the highly correlated measures of CD4 and CD19 levels colocalized at 18q21 (both P=.003). Similarly, a shared region of chromosome 1 was linked to CD8 and CD19 levels (P=.0001 and P=.002, respectively). Several of the identified chromosome regions are likely to harbor polymorphic candidate genes responsible for these important human phenotypes. Their discovery has important implications for understanding the generation of the immune repertoire and understanding immune-system homeostasis. More generally, these data show the power of an integrated human gene-mapping approach for heritable molecular phenotypes, using large pedigrees that have been extensively genotyped.

    View details for Web of Science ID 000175012400008

    View details for PubMedID 11951176

  • Natural killer cell immunoglobulin-like receptor (KIR) locus profiles in African and South Asian populations GENES AND IMMUNITY Norman, P. J., Carrington, C. V., Byng, M., Maxwell, L. D., Curran, M. D., Stephens, H. A., Chandanayingyong, D., Verity, D. H., Hameed, K., Ramdath, D. D., Vaughan, R. W. 2002; 3 (2): 86-95


    Natural killer (NK) and some T cells express killer cell immunoglobulin-like receptors (KIRs), which interact with HLA class I expressed by target cells and consequently regulate cytolytic activity. The number of KIR loci can vary and so a range of genetic profiles is observed. We have determined the KIR genetic profiles from one African (n = 62) and two South Asian (n = 108, n = 78) populations. Several of the KIRs are present at significantly different frequencies between the two major ethnic groups (eg KIR2DS4 gene frequency 0.82 African, 0.47 S Asian. Pc < 1 x 10(-6)) and this is due to uneven distribution of two KIR haplotype families 'A' and 'B'. All three populations described here displayed a greater degree of diversity of KIR genetic profiles than other populations investigated, which indicates further complexity of underlying haplotypes; in this respect we describe two individuals who appear homozygous for a large deletion including the previously ubiquitous 2DL4. We have also reanalysed three populations that we studied previously, for the presence of a KIR which is now known to be an indicator of the 'B' haplotype. South Asians had the highest overall frequencies of all KIR loci characteristic of 'B' haplotypes (Pc < 0.0001 to < 0.004). Furthermore, gene frequency independent deviances in the linkage disequilibrium were apparent between populations.

    View details for DOI 10.1038/sj/gene/6363836

    View details for Web of Science ID 000174646300004

    View details for PubMedID 11960306

  • Genetic determinism in the relationship between human CD4(+) and CD8(+) T lymphocyte populations? GENES AND IMMUNITY Ahmadi, K. R., Hall, M. A., Norman, P., Vaughan, R. W., Snieder, H., Spector, T. D., Lanchbury, J. S. 2001; 2 (7): 381-387


    The adaptive immune system in mammals acts in a coordinated manner to eliminate environmentally derived pathogens. Humans, mice and rats show within species variation in the levels and ratios of their peripheral CD4+ and CD8+ T cells and to a significant degree this variation is under the control of polymorphic genes. Whether genes act separately to specify CD4+ and CD8+ subpopulation levels or whether CD8+ variation is controlled through gene and environmental action on CD4+ cells or vice versa, is not known. We use a quantitative modelling approach in identical and non-identical female human twins to delineate the lines of control which act upon and between CD4+ and CD8+ subsets. The major findings of the study are: (1) genetic variation controls CD8+ T cell levels through two major routes-the first is via an effect on CD4+ T cells which accounts for the observed co-variation between CD4+ and CD8+ T cells, the second is through direct action on CD8+ T cell levels. (2) No evidence of a gene effect from CD8+ T cells on CD4+ cells is observed. Our findings have implications for the evolution of the complex defence system of which CD4+ and CD8+ T cells are a crucial part and encourage further work towards locating common pleiotropic quantitative trait loci responsible for variation in numbers of T cells.

    View details for Web of Science ID 000171980800005

    View details for PubMedID 11704804

  • Distribution of natural killer cell immunoglobulin-like receptor sequences in three ethnic groups IMMUNOGENETICS Norman, P. J., Stephens, H. A., Verity, D. H., Chandanayingyong, D., Vaughan, R. W. 2001; 52 (3-4): 195-205


    Killer cell immunoglobulin-like receptors (KIRs) are members of a group of molecules that specifically recognize HLA class I ligands and are found on subsets of human lymphopoetic cells. The number of KIR loci can vary between individuals, resulting in a heterogeneous array of possible KIR genes. The range of observed profiles has been explained by the occurrence of two haplotype families termed A and B which can be distinguished on the basis of certain KIR sequences. Here we attempted to determine whether the frequencies of putative KIR loci and the two haplotype groups vary in three ethnically defined, healthy, and unrelated control populations, namely UK Caucasoid (n=136), Palestinian (n=105) and Thai (n=119). We molecularly typed genomic DNA for the presence of 12 putative KIR loci, KIR2DL1, KIR2DL2, KIR2DL3, KIR2DL4, KIR3DL1, KIR3DL2, KIR2DS1, KIR2DS2, KIR2DS3, KIR2DS4, KIR2DS5, and KIR3DS1, using modified PCR sequence-specific primers. The patterns of KIR locus frequencies combined with the similar linkage disequilibrium values suggest that there was a distinction in the distribution of the two broad haplotype groups between the populations studied. The A haplotype was always the most prevalent, but the ratio of A to B varied between populations. The frequency of B haplotype was highest in the Palestinians and lowest in the Thais (Pc<0.0001).

    View details for Web of Science ID 000166746100005

    View details for PubMedID 11220621

  • Genetic influence on peripheral blood T lymphocyte levels GENES AND IMMUNITY Hall, M. A., Ahmadi, K. R., Norman, P., Snieder, H., MacGregor, A. J., Vaughan, R. W., Spector, T. D., Lanchbury, J. S. 2000; 1 (7): 423-427


    T lymphocytes are a major component of the adaptive immune system. CD4 positive T cell subpopulations regulate B cell and macrophage effector function while CD8 positive T cells are largely responsible for anti-viral cytotoxic activity. The degree of natural variation in the levels and ratios of the various T cell subpopulations is a possible risk factor for the development of autoimmune disease, infectious disease and cancer. There is some evidence from studies of inbred strains of mice and humans which suggests that variation in T cell subpopulations is genetically influenced. However, family studies alone cannot distinguish between common environmental and shared genetic influences and provide less robust estimates of the heritability than twin studies. To comprehensively examine genetic influences on a selection of important T cell phenotypes, we investigated variation in levels of total lymphocytes, CD3+, CD4+, CD8+, CD3+CD4+, CD3+CD8+ lymphocytes and in CD4:CD8 ratio as a proportion of lymphocytes and of T cells using the classical twin model approach. Healthy female twin pairs were sampled from the St. Thomas' UK Adult Twin Registry. A maximum of 103 monozygotic (MZ) and 186 dizygotic (DZ) twins aged 18-80 years participated in the study. Whole blood samples were analysed for T cell subsets by flow cytometry. The relative genetic contribution to these phenotypes was estimated using a variance components model-fitting approach. Heritability estimates were calculated of 65% for CD4:CD8 T cell and lymphocyte ratios, around 50% for absolute lymphocyte, CD3+ and CD4+ counts, and 56% for CD8+ numbers. Unique (rather than shared) familial environment explains the remainder of the variance. Genetic factors have a major influence on the variation in peripheral T cell subset numbers. Polymorphism dictating such variation should be taken into account when assessing risk factors for T cell immune-mediated disease with a genetic background.

    View details for Web of Science ID 000089770300003

    View details for PubMedID 11196672

  • PRESENCE OF IGG HLA ANTIBODIES IN PATIENTS WITH LONG-SURVIVING RENAL-ALLOGRAFTS XVth World Congress of the Transplantation-Society Norman, P. J., Harmer, A. W., Koffman, C. G., Vaughan, R. W. ELSEVIER SCIENCE INC. 1995: 999–1000

    View details for Web of Science ID A1995QJ19900394

    View details for PubMedID 7879260