Education & Certifications
Bachelor of Science, University of Washington, Biochemistry (2010)
Systematic screens for genetic interactions are a cornerstone of both network and systems biology. However, most screens have been limited to characterizing interaction networks in a single environment. Moving beyond this static view of the cell requires a major technological advance to increase the throughput and ease of replication in these assays. Here, we introduce iSeq-a platform to build large double barcode libraries and rapidly assay genetic interactions across environments. We use iSeq in yeast to measure fitness in three conditions of nearly 400 clonal strains, representing 45 possible single or double gene deletions, including multiple replicate strains per genotype. We show that iSeq fitness and interaction scores are highly reproducible for the same clonal strain across replicate cultures. However, consistent with previous work, we find that replicates with the same putative genotype have highly variable genetic interaction scores. By whole-genome sequencing 102 of our strains, we find that segregating variation and de novo mutations, including aneuploidy, occur frequently during strain construction, and can have large effects on genetic interaction scores. Additionally, we uncover several new environment-dependent genetic interactions, suggesting that barcode-based genetic interaction assays have the potential to significantly expand our knowledge of genetic interaction networks.
View details for DOI 10.1534/g3.116.034207
View details for PubMedID 27821633
Endogenous intestinal microbiota have wide-ranging and largely uncharacterized effects on host physiology. Here, we used reverse-phase liquid chromatography-coupled tandem mass spectrometry to define the mouse intestinal proteome in the stomach, jejunum, ileum, cecum and proximal colon under three colonization states: germ-free (GF), monocolonized with Bacteroides thetaiotaomicron and conventionally raised (CR). Our analysis revealed distinct proteomic abundance profiles along the gastrointestinal (GI) tract. Unsupervised clustering showed that host protein abundance primarily depended on GI location rather than colonization state and specific proteins and functions that defined these locations were identified by random forest classifications. K-means clustering of protein abundance across locations revealed substantial differences in host protein production between CR mice relative to GF and monocolonized mice. Finally, comparison with fecal proteomic data sets suggested that the identities of stool proteins are not biased to any region of the GI tract, but are substantially impacted by the microbiota in the distal colon.The ISME Journal advance online publication, 17 November 2015; doi:10.1038/ismej.2015.187.
View details for DOI 10.1038/ismej.2015.187
View details for Web of Science ID 000374377200014
View details for PubMedID 26574685
Transforming growth factor-β (TGF-β) signaling is required for normal vascular development. We aimed to discover the role of TGF-β signaling in embryonic smooth muscle cells (SMCs).We bred mice with smooth muscle (SM) 22α-Cre and Tgfbr2(flox) alleles to generate embryos in which the type II TGF-β receptor (TGFBR2; required for TGF-β signaling) was deleted in SMCs. Embryos were harvested between embryonic day (E) 9.5 and E18.5 and examined grossly, microscopically, and by histochemical and RNA analyses. SM22α-Cre(+/0) Tgfbr2(flox/flox) (knockout [KO]) embryos died before E15.5 with defects that included cardiac outflow tract abnormalities, persistence of the right dorsal aorta, and dilation of the distal aorta. Histological analyses suggested normal expression of SMC differentiation markers in KO aortas; however, RNA analyses showed that SMC differentiation markers were increased in KO cardiac outflow vessels but decreased in the descending aorta. KO aortas had only rare mature elastin deposits and contained abnormal aggregates of extracellular matrix proteins. Expression of several matrix proteins was significantly decreased in KO descending aortas but not in cardiac outflow vessels.TGF-β signaling in SMCs controls differentiation, matrix synthesis, and vascular morphogenesis. Effects of TGF-β on SMC gene expression appear to differ depending on the location of SMCs in the aorta.
View details for DOI 10.1161/ATVBAHA.111.238410
View details for Web of Science ID 000298288700001
View details for PubMedID 21979435
The mechanisms of atherosclerotic plaque rupture are poorly understood. Urokinase-type plasminogen activator (uPA) is expressed at elevated levels by macrophages in advanced human plaques. Patients with evidence of increased plasminogen activation have an elevated risk of major cardiovascular events. We used atherosclerotic mice to test the hypothesis that increased macrophage uPA expression in advanced plaques would cause histological features similar to those in ruptured human plaques.Bone marrow from transgenic mice with increased macrophage uPA expression or nontransgenic controls (all apolipoprotein E-null [Apoe(-/-)]) was transplanted into 35-week-old Apoe(-/-) recipients, and innominate lesions and aortas were examined 8 to 13 weeks later. Donor macrophages accumulated in innominate lesions adjacent to plaque caps and in aortas, increasing uPA expression at both sites. Recipients of uPA-overexpressing macrophages had an increased prevalence of intraplaque hemorrhage (61% versus 13%; P=0.002) as well as increased lesion fibrin staining and fibrous cap disruption (P=0.06 for both). Transplantation of uPA-overexpressing macrophages increased aortic matrix metalloproteinase activity (40%; P=0.02). This increase was independent of matrix metalloproteinase-9.In advanced plaques of Apoe(-/-) mice, macrophage uPA overexpression causes intraplaque hemorrhage and fibrous cap disruption, features associated with human plaque rupture. uPA overexpression also increases vascular matrix metalloproteinase activity. These data provide a mechanism that connects macrophage uPA expression, matrix metalloproteinase activity, and plaque rupture features in mice. The data also suggest that elevated plaque plasminogen activator expression and plasminogen activation in humans may be causally linked to plaque rupture and cardiovascular events.
View details for DOI 10.1161/CIRCULATIONAHA.109.914945
View details for Web of Science ID 000276530600010
View details for PubMedID 20351234
Impairment of transforming growth factor (TGF)-beta1 signaling accelerates atherosclerosis in experimental mice. However, it is uncertain whether increased TGF-beta1 expression would retard atherosclerosis. The role of TGF-beta1 in aneurysm formation is also controversial. We tested whether overexpression of active TGF-beta1 in hyperlipidemic mice affects atherogenesis and aortic dilation.We generated apolipoprotein E-null mice with transgenes that allow regulated overexpression of active TGF-beta1 in their hearts. Compared to littermate controls, these mice had elevated cardiac and plasma TGF-beta1, less aortic root atherosclerosis (P< or =0.002), fewer lesions in the thoracic and abdominal aortae (P< or =0.01), less aortic root dilation (P<0.001), and fewer pseudoaneurysms (P=0.02). Mechanistic studies revealed no effect of TGF-beta1 overexpression on plasma lipids or cytokines, or on peripheral lymphoid organ cells. However, aortae of TGF-beta1-overexpressing mice had fewer T-lymphocytes, more collagen, less lipid, lower expression of inflammatory cytokines and matrix metalloproteinase-13, and higher expression of tissue inhibitor of metalloproteinase-2.When overexpressed in the heart and plasma, TGF-beta1 is an antiatherogenic, vasculoprotective cytokine that limits atherosclerosis and prevents aortic dilation. These actions are associated with significant changes in cellularity, collagen and lipid accumulation, and gene expression in the artery wall.
View details for DOI 10.1161/ATVBAHA.109.186593
View details for Web of Science ID 000269098200003
View details for PubMedID 19325140