This Mentored Research Scientist Development Award (K01) application aims to expand upon the base knowledge I have acquired during my post-doctoral training in the genetics of diabetic nephropathy (DN) and develop expertise in both analytical and laboratory skills that will prepare me for a long and impactful career in research on the genetics of diabetes and its complications. Through carefully structured career development and training activities, an exceptional network of mentored support, and a research plan aimed at addressing specific challenges in investigations on the genetics of DN, I fully expect to accomplish this goal. Recent advances in population genetics and genomics have expanded our understanding of genetic variation across our genome and facilitated cutting-edge technologies that have transformed our ability to identify genes that underlie human disease. Genome-wide association (GWA) studies have spearheaded this revolution, successfully identifying hundreds of common genetic variants associated with complex disease. Despite these successes, a number of important challenges lie ahead to fully appreciate the genetic basis of these complex diseases. At the forefront of these challenges is pinpointing the causal functional variants that underlie the identified associations, a task that is particularly formidable when the associated SNP is localized to a region devoid of evidence of protein-coding genes. Additionally, somewhat surprisingly, the first wave of GWA studies has shown that the effect size of common variants is more modest than previously suspected, leading to speculation that other sources, including rare, low frequency variants, account for a portion of this 'missing heritability'. Each of these challenges was manifest in our recent GWA scan of DN in the Genetics of Kidneys in Diabetes (GoKinD) type 1 diabetes (T1D) Nephropathy Collection. With these challenges in mind, my proposed training seeks to build on my previous experience while moving this area of research forward. This will be accomplished through extensive didactic and hands-on training in both analytical and laboratory aspects of population genetics and genomics that includes the development of statistical and analytical skills, training in new methodologies of next-generation sequencing and gene regulation to investigate the relationship between genetic variations and disease predisposition, and training in analytical methodologies and research strategies for investigating complex disease. My proposed research aims to further investigate genetic associations identified in our GWA scan of the GoKinD collection on chromosomes 1p and 13q and identify both i) the causative genetics variants and ii) the disease genes underlying the associations at each of these loci. This goal will be accomplished through two Specific Aims. 1) To identify DNA sequence differences that are causally related to DN in the associated regions on chromosomes 1p and 13q. To accomplish this, I will a) establish a comprehensive catalog of all DNA sequence differences located in the intervals containing the associated SNPs using next-generation sequencing technology. This will be accomplished by targeted resequencing of these regions, including the haplotype blocks/LD intervals containing the associated variants and the exons of all known genes in the 1p and 13q regions, in pooled DNA samples from the GoKinD T1D Nephropathy Collection using filter-based hybridization capture followed by massively parallel DNA resequencing. I will then b) prioritize these candidate causal variants based on their bioinformatic, biological, and statistical significance. This will be accomplished using a variety of criteria to identify the most promising functional candidates followed by follow-up genotyping in the entire GoKinD T1D Nephropathy Collection. 2) To identify the DN susceptibility genes at the chromosome 1p and 13q regions by mapping the candidate causal variants to their target DN disease gene. To accomplish this I will a) establish a comprehensive renal transcript map for the associated regions on chromosome 1p and 13q using RNA-Seq technology and kidney biopsies obtained from T1D patients with early DN, b) map the long-range interaction between the candidate DN risk variants on 13q and its distal target gene using Chromosome Conformation Capture technology, and c) perform in vitro studies to examine the impact of each candidate causal variant on the expression of their target DN susceptibility gene within the 1p and 13q chromosomal regions. These targeted studies will be the foundation for future, more detailed functional studies. Together, this goal and these Specific Aims offer a tractable course of investigation that builds on convincing preliminary data. They will have a profound impact on our understanding of the allelic architecture of DN and its underlying mechanism. Importantly, they also faithfully integrate highly-focused research activities with my overall training and career development goals.
The proposed training aims to expand upon the base knowledge I have acquired during my post-doctoral training in the genetics of diabetic nephropathy (DN) and develop expertise in both analytical and laboratory skills that will prepare me for a long and impactful career in research on the genetics of diabetes and its complications. This will be accomplished through extensive didactic and hands-on training in both analytical and laboratory aspects of population genetics and genomics. The proposed research strategy will have a profound impact on our understanding of the allelic architecture of DN and its underlying mechanism while faithfully integrating highly-focused research activities with my overall training and career development goals.
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|Prudente, Sabrina; Shah, Hetal; Bailetti, Diego et al. (2015) Genetic Variant at the GLUL Locus Predicts All-Cause Mortality in Patients With Type 2 Diabetes. Diabetes 64:2658-63|
|Pezzolesi, Marcus G; Krolewski, Andrzej S (2013) The genetic risk of kidney disease in type 2 diabetes. Med Clin North Am 97:91-107|
|Pezzolesi, Marcus G; Jeong, Jackson; Smiles, Adam M et al. (2013) Family-based association analysis confirms the role of the chromosome 9q21.32 locus in the susceptibility of diabetic nephropathy. PLoS One 8:e60301|
|Martini, Sebastian; Nair, Viji; Patel, Sanjeevkumar R et al. (2013) From single nucleotide polymorphism to transcriptional mechanism: a model for FRMD3 in diabetic nephropathy. Diabetes 62:2605-12|