Anne E Kwitek
University Of Iowa
Project start date: 2008-06-01
Project end date: 2013-04-30
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Grants awarded to Anne E Kwitek
Application Of Genetics And Physiological Genomics To Dissect Resistance To T1D
Anne E Kwitek, Assistant Professor
Medical College Of Wisconsin
8701 Watertown Plank Rd
milwaukee, Wi 532260509
Grant 5P01AI042380-120006 from National Institute Of Allergy And Infectious Diseases IRG: ZAI1
Abstract: Type 1 diabetes (T1D) is one of the prototypical organ-specific autoimmune endocrinopathies that results in life-long dependency on daily insulin injection. Molecular genetic approaches have identified regions of the human, mouse, rat genomes that play a role in susceptibility to Type 1 Diabetes (T1D). While the major histocompatibility complex (MHC), the equivalent of the human leukocyte antigen (HLA), is the major locus responsible for susceptibility to T1D, there is a separate set of genes that contribute to susceptibility and end organ damage. We have been studying the genetic factors contributing to T1D in the BBDP rat, a model of spontaneous disease concurrent with lymphopenia. We have identified the location of five loci involved in T1D Iddm2/lyp, lddm1/MHC, Iddm3, Iddm19, and a fifth locus on chromosome 4 (Iddm4?). Given the identification of the Ian5 mutation as the gene responsible for Iddm2/lyp, we now focus on the identification of the additional factors playing a role in T1D in the BB rat. Our mapping studies have identified four diabetogenic loci in addition to the MHC, which Projects 3 specifically addresses, in close collaboration with Project 1. The focus of Project 3 is to generate consomic and derived congenic strains for three additional factors, including Iddm3, Iddm19, and the susceptibility locus on chromosome 4 nearby Ian5 (Project 1) and to evaluate these new models for their role in T1D using gene expression profiling, comparative genomics, and sequence analysis. Furthermore, we are developing the technology to knock genes out in the rat for validation of positionally cloned genes. Specifically we will 1. Continue positional cloning efforts for Iddm3 to identify sequence variants contributing to resistance to autoimmune diabetes. 2. Follow up a second resistance factor on chromosome 15. 3. Use novel 70-mer oligonucleotide and cDNA microarray platforms to evaluate genes and pathways affected by the diabetogenic factors identified in the BB rat. 4. Generate rat ENU-induced knockout strains to validate genes involved in autoimmune diabetes in the BB rat
Keywords: diabetes mellitus genetics, functional /structural genomics, insulin dependent diabetes mellitus gene expression, genetic mapping, genetic susceptibility, lymphopenia, major histocompatibility complex gene expression profiling, genetically modified animal, laboratory rat, microarray technology
DISSECTING THE GENETICS OF THE METABOLIC SYNDROME ON CHROMOSOME 17 OF THE LH RAT
Anne E Kwitek
University Of Iowa, Iowa City, Ia 52242
Grant 5R01HL089895-03 from National Heart, Lung, And Blood Institute
Abstract: The collection of maladies including obesity, dyslipidemia, insulin resistance and hypertension is referred to as the metabolic syndrome and is a major risk factor for cardiovascular disease. Its incidence continues to rise, in part as a result of the epidemic increase in obesity. The Lyon Hypertensive (LH) rat is a powerful inbred model for dissecting the genetic contributions of the human metabolic syndrome, with high body weight, cholesterol, triglycerides, and insulin/glucose ratios, and salt-sensitive hypertension. Interestingly, the Lyon normotensive (LN) control strain, concurrently selected from the same SD colony, is genetically quite similar (85%) to the LH, but phenotypically very distinct. Genetic linkage mapping in an LH x LN intercross identified 3 clusters of Quantitative Traits Loci (QTL) on rat chromosome (RNO) 17 for multiple features of the metabolic syndrome in the LH rat and 3 QTL for blood pressure on RNO17, 2, and 13. We hypothesize that the LH chromosome 17 has genes contributing to all major traits of the metabolic syndrome, and that there are QTL interactions between QTL on RNO 17 and between RNO 17, 2, and 13. To identify at least one of these genes, we propose a combined traditional positional cloning, in silico mapping, and QTL (phenotype and gene expression) mapping to identify causal genes and pathways leading to the metabolic syndrome. We will 1 Validate the QTL regions for RNO17 in the LH and LN strains. The close genetic identity of the LH and LN strains allows for rapid generation and characterization of a consomic LH-17LN strain and congenic substrains. These strains will be used to narrow the QTL regions and to determine which phenotypes are due to multiple QTL, interacting QTL, or pleiotropic effects of a single gene within a QTL. 2 Investigate interactions between trans-acting and cis-acting haplotypes on traits underlying the metabolic syndrome. We will perform an F2 intercross between LH and LN to determine the interactions between QTL on RNO17, 2, and 13, and regulatory networks causing the phenotypes we observe in the LH. The offspring will be comprehensively studied by phenotyping, SNP genotyping, and gene expression profiling for combined phenotype (p) QTL and expression (e) QTL mapping. 3 Refine the QTL on RNO17 through SNP fine-mapping and experimental validation in congenic rats. For each candidate region, we will fine-map the haplotypes by additional SNP typing to narrow and define minimal QTL intervals. All genetic variation will be determined in these minimal regions to identify candidate genes and sequence variants. These regions will be experimentally validated in the congenics. 4 Validate functional variants through rat transgenic rescue. For the strongest candidate with evidence of functional sequence variants identified in the above aims, we will develop a transgenic rat, either introducing the LN allele into the LH rat or the LH allele into the LH-17LN
Keywords: Affect; Alleles; Allelomorphs; Blood Pressure; Blood Pressure, High; Body Weight; Candidate Disease Gene; Candidate Gene; Cardiovascular Diseases; Cessation of life; Cholest-5-en-3-ol (3beta)-; Cholesterol; Chromosome 1; Chromosome 17; Chromosome 2; Chromosome Mapping; Chromosomes; Chromosomes, Human; Chromosomes, Human, Pair 1; Chromosomes, Human, Pair 17; Chromosomes, Human, Pair 2; Collection; Common Rat Strains; Complex; Computer Simulation; Computerized Models; Congenic Strain; D-Glucose; DNA Resequencing; Death; Dextrose; Diet; Disease; Disorder; Dyslipidemias; Epidemic; Gene Combinations; Gene Expression; Gene Expression Monitoring; Gene Expression Pattern Analysis; Gene Expression Profiling; Gene Localization; Gene Mapping; Gene Mapping, Total Human and Non-Human; Gene variant; Generations; Genes; Genes, Regulator; Genetic; Genetic Diversity; Genetic Identity; Genetic Variation; Genetics, Gene Mapping; Genome; Genomics; Glucose; Haplotypes; Human; Human Chromosomes; Human, General; Humulin R; Hyperglyceridemia; Hypertension; Hypertriglyceridemia; Inbred Strain; Incidence; Insulin; Insulin (ox), 8A-L-threonine-10A-L-isoleucine-30B-L-threonine-; Insulin Resistance; Insulin, Regular; Lead; Link; Linkage (Genetics); Linkage Mapping; Lipids; Mammals, Rats; Man (Taxonomy); Man, Modern; Maps; Mathematical Model Simulation; Mathematical Models and Simulations; Metabolic syndrome; Modeling; Models, Computer; Novolin R; Obesity; Organ failure; Pathway interactions; Pb element; Phenotype; Profilings, Gene Expression; QTL; Quantitative Trait Loci; Raised TG; Raised triglycerides; Rat; Rattus; Regulator Genes; Research; Research Resources; Resequencing; Resistance; Resources; Risk Factors; Role; SNP genotyping; Schools, Medical; Simulation, Computer based; Sodium Chloride; Sodium chloride (NaCl); Trans-Acting Factors; Trans-Activators; Transactivators; Transcript Expression Analyses; Transcript Expression Analysis; Transcriptional Regulatory Elements; Transgenic Organisms; Triacylglycerol; Triglyceride increased; Triglycerides; Triglycerides high; Validation; Variant; Variation; Variation (Genetics); Vascular Hypertensive Disease; Vascular Hypertensive Disorder; Wisconsin; adiposity; allelic variant; cardiovascular disorder; computational modeling; computational models; computational simulation; computer based models; computerized modeling; computerized simulation; congenic; consomic; corpulence; corpulency; corpulentia; disease/disorder; elevated tg; elevated triglyceride; gene interaction; genetic linkage; genetic mapping; heavy metal Pb; heavy metal lead; hyperpiesia; hyperpiesis; hypertensive disease; in silico; insulin resistant; medical schools; normotensive; obese; obese people; obese person; obese population; offspring; pathway; positional cloning; regulatory gene; resistant; resistant strain; reverse genetics; salt; salt sensitive; social role; trait; trans acting element; trans acting factor (genetic); transgenic; virtual simulation
Project start date: 2008-06-01
Project end date: 2012-04-30
Budget start date: 1-MAY-2010
Budget end date: 30-APR-2011
PFA/PA: PA-07-070
5R01HL089895-03 (2010): $375808
5R01HL089895-02 (2009): $375688
Application Of Genetics And Physiological Genomics To Dissect Resistance To T1D
Anne E Kwitek, Assistant Professor
Medical College Of Wisconsin 8701 Watertown Plank Rd Milwaukee, Wi 532260509
Grant 5P01AI042380-110006 from National Institute Of Allergy And Infectious Diseases IRG: ZAI1