Biology subjects

Chan, K.

Publications and source records attributed to Chan, K..

4 recordsLinked to original sources

bioRxiv2018

Process-specific somatic mutation distributions vary with three-dimensional genome structure

Somatic mutations arise during the life history of a cell. Mutations occurring in cancer driver genes may ultimately lead to the development of clinically detectable disease. Nascent cancer lineages continue to acquire somatic mutations throughout the neoplastic process and during cancer evolution (Martincorena and Campbell, 2015). Extrinsic and endogenous mutagenic factors contribute to the accumulation of these somatic mutations (Zhang and Pellman, 2015). Understanding the underlying factors generating somatic mutations is crucial for developing potential preventive, therapeutic and clinical decisions. Earlier studies have revealed that DNA replication timing (Stamatoyannopoulos et al., 2009) and chromatin modifications (Schuster-Bockler and Lehner, 2012) are associated with variations in mutational density. What is unclear from these early studies, however, is whether all extrinsic and exogenous factors that drive somatic mutational processes share a similar relationship with chromatin state and structure. In order to understand the interplay between spatial genome organization and specific individual mutational processes, we report here a study of 3000 tumor-normal pair whole genome datasets from more than 40 different human cancer types. Our analyses revealed that different mutational processes lead to distinct somatic mutation distributions between chromatin folding domains. APOBEC- or MSI-related mutations are enriched in transcriptionally-active domains while mutations occurring due to tobacco-smoke, ultraviolet (UV) light exposure or a signature of unknown aetiology (signature 17) enrich predominantly in transcriptionally-inactive domains. Active mutational processes dictate the mutation distributions in cancer genomes, and we show that mutational distributions shift during cancer evolution upon mutational processes switch. Moreover, a dramatic instance of extreme chromatin structure in humans, that of the unique folding pattern of the inactive X-chromosome leads to distinct somatic mutation distribution on X chromosome in females compared to males in various cancer types. Overall, the interplay between three-dimensional genome organization and active mutational processes has a substantial influence on the large-scale mutation rate variations observed in human cancer.

genomics↗

bioRxiv2018

Different capsid-binding patterns of the β-herpesvirus-specific tegument protein pp150 (pM32/pUL32) in murine and human cytomegaloviruses

The phosphoprotein pp150 is a structurally, immunogenically, and regulatorily important capsid-associated tegument protein abundant in {beta}-herpesviruses including cytomegaloviruses (CMV), but absent in -herpesviruses and {Gamma}-herpesviruses. In human CMV (HCMV), bridging across each triplex and three adjacent major capsid proteins (MCPs) is a group of three pp150 subunits in a \"{bigtriangleup}\"-shaped fortifying configuration, 320 of which encase and stabilize the genome-containing capsid. Because murine CMV (MCMV) has been used as a model for HCMV pathogenesis and therapeutic studies, one might expect that pp150 and the capsid in MCMV and HCMV have similar structures. Here, by cryoEM and sub-particle reconstructions, we have obtained structures of MCMV capsid and pp150 at near atomic resolutions and built their atomic models. Surprisingly, the capsid-binding patterns of pp150 differ between HCMV and MCMV despite their highly similar capsid structures. In MCMV, pp150 is absent on triplex Tc and exists as a \"{Lambda}\"-shaped dimer on other triplexes, leading to only 260 groups of two pp150 subunits per capsid in contrast to 320 groups of three pp150 subunits encasing each HCMV capsid. Many more amino acids contribute to pp150-pp150 interactions in MCMV than in HCMV, making MCMV pp150 dimer inflexible thus incompatible to instigate triplex Tc-binding as observed in HCMV. While pp150 is essential in HCMV, pp150-deleted MCMV mutants remained viable though with attenuated infectivity and exhibiting defects in retaining viral genome. These results support targeting capsid proteins, but invalidate targeting pp150, when using MCMV as a model for HCMV pathogenesis and therapeutic studies.\n\nImportanceCMV infection is a leading viral cause of congenital birth defects and often responsible for life-threating complications in immunocompromised individuals like AIDS and post-organ transplantation patients. Absence of effective vaccines and potent drugs against CMV infections has motivated animal-based studies, mostly based on the mouse model with MCMV, both for understanding pathogenesis of CMV infections and for developing therapeutic strategies. Here, we present the first atomic structures of MCMV and show that the organization patterns of capsid-associated tegument protein pp150 between human and mouse CMV are different despite their highly similar capsid structures. Our functional studies demonstrate that deleting pp150 does not eliminate MCMV infection in contrast to pp150s essential role in HCMV infections. These results thus establish the validity to target capsid proteins, but raise concerns to target pp150, when using MCMV as HCMV model for pathogenesis and therapeutic studies.

microbiology↗

bioRxiv2017

Maternal obesity alters uterine NK cell activity through a functional KIR2DL1/S1 imbalance

In pregnancy, uterine natural killer cells (uNK) play essential roles in coordinating uterine angiogenesis, blood vessel remodeling, and promoting maternal tolerance to fetal tissue. Deviances from a normal uterine microenvironment are thought to modify uNK function(s), limiting their ability to establish a healthy pregnancy. While maternal obesity has become a major health concern due to associations with adverse effects on fetal and maternal health, our understanding into how obesity contributes to poor pregnancy disorders is essentially unknown. Given the importance of uNK in pregnancy, this study sets out to examine if obesity affects uNK function. Using a cohort of pregnant women, we show that baseline activity of uNK from obese women is elevated, but that enhanced activity does not equate to increased killing potential. Instead, obesity associates with altered uNK production of angiogenic VEGF-A and PlGF. These changes coincide with alterations in NKp46+ and NKG2A+ uNK subsets and elevated expression of KIR2D(L1/S1/S3/S5) receptors. Detailed examination revealed that obesity leads to imbalances in KIR2DL1/S1 expression that together instruct altered responses to HLA-C2 antigen, including increased production of TNF. Together, these findings suggest that maternal obesity modulates uNK function by altering angiokine/cytokine production and the response to HLA-C2 antigen.

immunology↗

bioRxiv2017

Evaluation and Design of Genome-wide CRISPR/Cas9 Knockout Screens

The adaptation of CRISPR/Cas9 technology to mammalian cell lines is transforming the study of human functional genomics. Pooled libraries of CRISPR guide RNAs (gRNAs), targeting human protein-coding genes and encoded in viral vectors, have been used to systematically create gene knockouts in a variety of human cancer and immortalized cell lines, in an effort to identify whether these knockouts cause cellular fitness defects. Previous work has shown that CRISPR screens are more sensitive and specific than pooled library shRNA screens in similar assays, but currently there exists significant variability across CRISPR library designs and experimental protocols. In this study, we re-analyze 17 genome-scale knockout screens in human cell lines from three research groups using three different genome-scale gRNA libraries, using the Bayesian Analysis of Gene Essentiality (BAGEL) algorithm to identify essential genes, to refine and expand our previously defined set of human core essential genes, from 360 to 684 genes. We use this expanded set of reference Core Essential Genes (CEG2), plus empirical data from six CRISPR knockout screens, to guide the design of a sequence-optimized gRNA library, the Toronto KnockOut version 3.0 (TKOv3) library. We demonstrate the high effectiveness of the library relative to reference sets of essential and nonessential genes as well as other screens using similar approaches. The optimized TKOv3 library, combined with the CEG2 reference set, provide an efficient, highly optimized platform for performing and assessing gene knockout screens in human cell lines.

systems biology↗