Despite decades of progress in unraveling cellular and molecular regulatory mechanisms that protect against autoimmunity, we still have only a marginal understanding of how lymphocytes "break the rules" in escaping these self-tolerance mechanisms to participate in autoimmune disease. Nearly all investigations of disease etiology have focused on familial genetics and environmental triggers that range from infections and xenobiotic insults to vitamin deficiencies. Yet none of these have provided sufficient mechanistic explanation for the stochastic nature of autoimmunity, which is still treated with relatively crude and often ineffective therapies. In this project, we propose to develop a model to test a largely ignored third alternative. We will test the hypothesis that somatic mutagenesis in lymphocytes is a major contributor to autoimmune disease. Though many lines of evidence point to this idea, there has been a dearth of associated study. The reason is simple: to date the hypothesis has been largely intractable to investigation. However, recent technical advances in DNA sequencing and stem cell biology/regenerative medicine have converged to open a heretofore locked door. We can now address the mutation/autoimmunity hypothesis in the context of the nonobese diabetic mouse (NOD) model of type I diabetes. Our laboratory is in the unique position of having direct access to a large panel of diabetogenic CD4 T cell clones derived from splenocytes of diabetic NOD mice. In two aims, we will make use of these clones to test the hypothesis that they have been functionally altered by somatic mutations, enabling their escape from self-tolerance and promoting their pancreatic islet-specific pathology. We will use NextGen sequencing to uncover somatic mutations in the exomes of diabetogenic CD4 T cell clones and perform bioinformatic pathways analyses to identify signaling pathways affected by mutations. In a complementary aim, we will evaluate somatic mutations in the entire genome of one clone, BDC-2.5, by generating induced pluripotent stem cells (iPSC) in order to erase the epigenetic program of the BDC-2.5 clone. CD4 T cells subsequently derived from BDC-2.5 iPSC will be functionally compared head-to-head with CD4 T cells of transgenic mice that share the BDC-2.5 clone ??T cell receptor but not the BDC-2.5 clone- specific somatic mutations. In the context of adoptive transfer and bone marrow chimera studies, this comparison will enable us to determine the effects of genome-wide somatic mutations on pathological properties of the BDC-2.5 clone. Uncovering a role for somatic mutagenesis in diabetes would transform investigation of this and other autoimmune diseases, opening new avenues to identify targets for rational therapeutic intervention. Regardless of outcome, our study will be the first to directly assess the contributions of somatic genetics versus epigenetics to autoimmunity.

Public Health Relevance

This is a proposal to uncover the molecular mechanisms that promote autoimmune disease. We will test the idea that, as in cancer, somatic mutations in DNA are responsible for the pathological properties of rogue lymphocytes that initiate and mediate autoimmunity. This is a game-changing hypothesis because prior studies have focused almost entirely on familial genetics and environmental triggers as causative factors in disease. Testing the somatic mutation idea has been intractable until now. However, circumstance together with a number of technological advances have recently converged, placing us in a unique position to test the somatic mutation/autoimmunity idea in context of a mouse model of type I diabetes. Although our study focuses on diabetes, its basic nature will likely have a far-reaching impact on how many autoimmune diseases are investigated and treated.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21AI113122-01
Application #
8762869
Study Section
Special Emphasis Panel (ZRG1-HAI-K (08))
Program Officer
Bourcier, Katarzyna
Project Start
2014-06-01
Project End
2016-05-31
Budget Start
2014-06-01
Budget End
2015-05-31
Support Year
1
Fiscal Year
2014
Total Cost
$247,025
Indirect Cost
$91,173
Name
National Jewish Health
Department
Type
DUNS #
076443019
City
Denver
State
CO
Country
United States
Zip Code
80206