Browsing by Subject "GWAS"

Genetics account for a large, mostly unexplained proportion of human disease. Though the role of genetics in simple, Mendelian traits has long been established, it is more difficult to disambiguate the role of various human genetic factors in complex disease traits. However, as genetics technology and methodology has advanced, from genome-wide association studies (GWAS) to next-generation sequencing (NGS), our ability to detect the role of both rare and common human genetic variation in complex disease traits has greatly improved, allowing us to demonstrate robust genetic factors involved in a variety of disease from metabolic to viral. However, despite the outstanding progress in human genetics, many complex disease traits lack robustly associated genetic variants, the existing variation only accounts for a small proportion of the estimated heritability, or the trait lacks comprehensive genetic investigation all together.

In this thesis I conducted a common variant study using GWAS and a comprehensive NGS analysis - both standards in the field - to investigate the role of human genetics in the severity of complex disease traits ranging from viral disease to metabolic: herpes simplex virus type 2 (HSV-2) and non-alcoholic fatty liver disease (NAFLD). Chapter 1 provides a broad overview of current human genetics methodologies and the advantages and caveats to each technology for complex disease traits, as well as the background and current state of genetics research for the two complex traits investigated: HSV-2 and NAFLD.

Chapter 2 utilizes a GWAS to investigate the role of common human genetic variation in HSV-2 severity, which has previously only been investigated through a small handful of candidate gene studies. We were unable to replicate previous candidate gene associations, though we did detect several variants in or near biologically plausible genes (including ABCA1 and KIF1B) that approached, though did not reach, genome-wide statistical significance with HSV-2 severity as measured by the quantitative viral shedding rate. This is the first genome-wide investigation of human genetics in HSV-2.

Chapter 3 utilizes whole-exome sequencing at both the single-variant and gene levels to further elucidate the role of human genetics in gold standard liver biopsy confirmed NAFLD fibrosis extreme phenotypes: protective and progressor. We were able to replicate known associations with PNPLA3 and TM6SF2 and advanced fibrosis, despite the limited available sample size. We also observed enrichment of variation in distinct genes for progressor or protective NAFLD phenotypes, though these genes did not reach statistical significance. This is the first NGS study of NAFLD, and thus the first investigation of the role of rare variation in NAFLD.

Overall, this thesis applied genome-wide techniques to interrogate gaps in the genetics of complex trait severity, from viral to liver disease, using unique, well-phenotyped cohorts. Human genetics remains a complicated field that will require the continued use of well-phenotyped cohorts in larger numbers, as well as both complementary and confirmatory sequencing and bioinformatics methods to fully detangle. While the research in this thesis is primarily hypothesis generating, and potentially associated variants will have to be replicated and investigated on a functional level to be confirmed as causal, the exploration of genetic associations with complex disease traits can prove highly informative for both understanding the underlying biology of these traits and for identifying genes and pathways that may act as biomarkers or treatment targets. Thus, this thesis has acted as a primer to expand knowledge of the role of human genetics in two highly complex and varied traits, HSV-2 and NAFLD, paving the way for further studies, ultimately with the goal of improving human health.

The maintenance of genetic variation for quantitative traits has long puzzled evolutionary biologists. Previous studies have effectively interrogated large genomic regions both in the greenhouse and field to understand the selective forces maintaining variation. However, subtler points can be difficult to tease out from these genomic regions. Here, we take advantage of advances in sequencing technology to understand how variation for flowering time and size is maintained within a single population of Mimulus guttatus at the nucleotide level. By focusing on the nucleotide level, we directly address questions of allelic effect and frequency, and interrogate polymorphisms found to be significantly associated with traits in the greenhouse in other contexts to determine their contributions to growth and fitness.

Flowering time and flower size have been shown to be highly quantitative traits within the Iron Mountain population of M. guttatus, and it has been proposed that minor alleles are likely to increase trait values. We present here confirmation of this long-standing hypothesis. By creating a genome wide association (GWA) mapping panel, we have identified multiple significant site-trait associations within the IM population of M. guttatus, and found extensive evidence of pleiotropy and polygenic adaptation.

To understand how developmental processes might contribute to variation in flowering time and flower size, we next investigate genetic variation for circadian period length. The circadian clock is known to be upstream of floral induction pathways, and in Arabidopsis thaliana over one third of the genome is suspected to be under circadian regulation. We find extensive variation for endogenous period length, and genetic correlations between endogenous period length and days to germination, flower size measurements, height, and leaf size. Despite a small GWA analysis sample size, we find abundant evidence of overlap of sites found to be significant for growth and flowering traits and period length.

Finally, we investigate how genomic variants identified in the greenhouse contribute to variation in a natural context. By planting F1 crosses of the lines sequenced for the GWA mapping panel in the field, we are able to confirm significant impacts on field growth and fitness, and identify a genomic site significantly associated with survival to seed production. Taken together our results not only confirm previous quantitative trait locus work at the nucleotide level, but present a newly developed sequence resource for analysis of intra-population variation in M. guttatus.