Doctor of Philosophy, California Institute of Technology (2009)
William Talbot, Postdoctoral Faculty Sponsor
Microglia are phagocytic cells that form the basis of the brain's immune system. They derive from primitive macrophages that migrate into the brain during embryogenesis, but the genetic control of microglial development remains elusive. Starting with a genetic screen in zebrafish, we show that the noncanonical NOD-like receptor (NLR) nlrc3-like is essential for microglial formation. Although most NLRs trigger inflammatory signaling, nlrc3-like acts cell autonomously in microglia precursor cells to suppress unwarranted inflammation in the absence of overt immune challenge. In nlrc3-like mutants, primitive macrophages initiate a systemic inflammatory response with increased proinflammatory cytokines and actively aggregate instead of migrating into the brain to form microglia. NLRC3-like requires both its pyrin and NACHT domains, and it can bind the inflammasome component apoptosis-associated speck-like protein. Our studies suggest that NLRC3-like may regulate the inflammasome and other inflammatory pathways. Together, these results demonstrate that NLRC3-like prevents inappropriate macrophage activation, thereby allowing normal microglial development.
View details for DOI 10.1016/j.celrep.2013.11.004
View details for Web of Science ID 000328266400017
View details for PubMedID 24316075
The vertebrate peripheral nervous system contains sensory neurons that arise from ectodermal placodes. Placodal cells ingress to move inside the head to form sensory neurons of the cranial ganglia. To date, however, the process of placodal cell ingression and underlying cellular behavior are poorly understood as studies have relied upon static analyses on fixed tissues. Visualizing placodal cell behavior requires an ability to distinguish the surface ectoderm from the underlying mesenchyme. This necessitates high resolution imaging along the z-plane which is difficult to accomplish in whole embryos. To address this issue, we have developed an imaging system using cranial slices that allows direct visualization of placode formation.We demonstrate an effective imaging assay for capturing placode development at single cell resolution using chick embryonic tissue ex vivo. This provides the first time-lapse imaging of mitoses in the trigeminal placodal ectoderm, ingression, and intercellular contacts of placodal cells. Cell divisions with varied orientations were found in the placodal ectoderm all along the apical-basal axis. Placodal cells initially have short cytoplasmic processes during ingression as young neurons and mature over time to elaborate long axonal processes in the mesenchyme. Interestingly, the time-lapse imaging data reveal that these delaminating placodal neurons begin ingression early on from within the ectoderm, where they start to move and continue on to exit as individual or strings of neurons through common openings on the basal side of the epithelium. Furthermore, dynamic intercellular contacts are abundant among the delaminating placodal neurons, between these and the already delaminated cells, as well as among cells in the forming ganglion.This new imaging assay provides a powerful method to analyze directly development of placode-derived sensory neurons and subsequent ganglia formation for the first time in amniotes. Viewing placode development in a head cross-section provides a vantage point from which it is possible to study comprehensive events in placode formation, from differentiation, cell ingression to ganglion assembly. Understanding how placodal neurons form may reveal a new mechanism of neurogenesis distinct from that in the central nervous system and provide new insight into how cells acquire motility from a stationary epithelial cell type.
View details for DOI 10.1186/1471-2202-12-37
View details for Web of Science ID 000291750600001
View details for PubMedID 21554727
Glypicans are conserved cell surface heparan sulfate proteoglycans expressed in a spatiotemporally regulated manner in many developing tissues including the nervous system. Here, we show that Glypican-1 (GPC1) is expressed by trigeminal placode cells as they ingress and contribute to trigeminal sensory neurons in the chick embryo. Either expression of full-length or truncated GPC1 in vivo causes defects in trigeminal gangliogenesis in a manner that requires heparan sulfate side chains. This leads to either abnormal placodal differentiation or organization, respectively, with near complete loss of the ophthalmic (OpV) trigeminal ganglion in the most severe cases after overexpression of full-length GPC1. Interestingly, modulating GPC1 alters levels of endogenous Wnt signaling activity in the forming trigeminal ganglion, as indicated by Wnt reporter expression. Accordingly, GPC1 overexpression phenocopies Wnt inhibition in causing loss of OpV placodal neurons. Furthermore, increased Wnt activity rescues the effects of GPC1 overexpression. Taken together, these results suggest that appropriate levels of GPC1 are essential for proper regulation of canonical Wnt signaling during differentiation and organization of trigeminal placodal cells into ganglia.
View details for DOI 10.1016/j.ydbio.2010.09.017
View details for Web of Science ID 000284438400013
View details for PubMedID 20883685
Vertebrate cranial sensory ganglia have a dual origin from the neural crest and ectodermal placodes. In the largest of these, the trigeminal ganglion, Slit1-Robo2 signaling is essential for proper ganglion assembly. Here, we demonstrate a crucial role for the cell adhesion molecule N-cadherin and its interaction with Slit1-Robo2 during gangliogenesis in vivo. A common feature of chick trigeminal and epibranchial ganglia is the expression of N-cadherin and Robo2 on placodal neurons and Slit1 on neural crest cells. Interestingly, N-cadherin localizes to intercellular adherens junctions between placodal neurons during ganglion assembly. Depletion of N-cadherin causes loss of proper ganglion coalescence, similar to that observed after loss of Robo2, suggesting that the two pathways might intersect. Consistent with this possibility, blocking or augmenting Slit-Robo signaling modulates N-cadherin protein expression on the placodal cell surface concomitant with alteration in placodal adhesion. Lack of an apparent change in total N-cadherin mRNA or protein levels suggests post-translational regulation. Co-expression of N-cadherin with dominant-negative Robo abrogates the Robo2 loss-of-function phenotype of dispersed ganglia, whereas loss of N-cadherin reverses the aberrant aggregation induced by increased Slit-Robo expression. Our study suggests a novel mechanism whereby N-cadherin acts in concert with Slit-Robo signaling in mediating the placodal cell adhesion required for proper gangliogenesis.
View details for DOI 10.1242/dev.034355
View details for Web of Science ID 000272076700009
View details for PubMedID 19934013
Vertebrate cranial sensory ganglia, responsible for sensation of touch, taste and pain in the face and viscera, are composed of both ectodermal placode and neural crest cells. The cellular and molecular interactions allowing generation of complex ganglia remain unknown. Here, we show that proper formation of the trigeminal ganglion, the largest of the cranial ganglia, relies on reciprocal interactions between placode and neural crest cells in chick, as removal of either population resulted in severe defects. We demonstrate that ingressing placode cells express the Robo2 receptor and early migrating cranial neural crest cells express its cognate ligand Slit1. Perturbation of this receptor-ligand interaction by blocking Robo2 function or depleting either Robo2 or Slit1 using RNA interference disrupted proper ganglion formation. The resultant disorganization mimics the effects of neural crest ablation. Thus, our data reveal a novel and essential role for Robo2-Slit1 signaling in mediating neural crest-placode interactions during trigeminal gangliogenesis.
View details for DOI 10.1038/nn2051
View details for Web of Science ID 000253548300009
View details for PubMedID 18278043
Cranial ectodermal placodes are critical for normal development of the peripheral nervous system of the head. However, many aspects of the molecular and tissue interactions involved in their induction have yet to be elucidated. The trigeminal placode is induced by an unidentified secreted factor(s) from the dorsal neural tube. To determine candidates that may be involved in this induction process, we have performed reverse transcriptase-polymerase chain reaction (RT-PCR) and whole-mount in situ hybridization to screen for receptors expressed by uninduced presumptive trigeminal level ectoderm. We have found that receptors for fibroblast growth factors, insulin-like growth factors, platelet-derived growth factors, Sonic hedgehog, the transforming growth factor-beta superfamily, and Wnts all are expressed in patterns consistent with a role in trigeminal placode formation. This RT-PCR screen for candidate receptors expressed in presumptive trigeminal ectoderm is the first systematic screen to identify potential interactions underlying induction of the trigeminal placode and represents a critical step for understanding this complex process.
View details for DOI 10.1002/dvdy.21325
View details for Web of Science ID 000250192100022
View details for PubMedID 17879314