HUMAN HYPOTHALAMIC NEURONAL EPIGENOMICS AND RISK FOR OBESITY

Translating information on genetic risk for body weight regulation into molecular mechanisms can have asignificant impact on intervention and therapies. We are seeking to identify genetic variation and theirmolecular mechanisms that influences obesity through direct effects on the hypothalamus as it is the brain hubthat regulates energy homeostasis and there is now considerable evidence for genetic influence to impact thisbrain region. However, the majority of genetic loci associated with this common, chronic disorder in the generalpopulation are located in noncoding regions of the genome, suggesting their influence on energy homeostasisis manifested through changes to the regulome. Thus, pinpointing the causal human variants and connectingthem to their downstream targets in brain presents challenges of tissue access for study because muchepigenetic control is species-, tissue- and context-specific. To overcome the barrier of limited human tissueaccess, we have developed a robust protocol for generating human induced pluripotent stem cell (iPSC)-derived neuronal cultures that recapitulate many of the features of hypothalamic neurons from the arcuatenucleus, including by benchmarking this in vitro model to in vivo events that are pivotal in hypothalamicdevelopment. We will use this human model and state of the art high throughput assays to map the currentlyuncharted regulatory landscape of the human hypothalamic neurons across 3 stages in development (early,mid, and late) and under experimental obesogenic conditions. Next, in order to precisely pinpoint the functionalvariants in BMI GWAS loci that have influence on body weight regulation through hypothalamic epigenomicregulation, we will identify those that influence chromatin accessibility and/or target gene expression by assayin 100 iPSC-derived neuronal lines generated from subjects of the San Antonio Mexican American FamilyStudies. GWAS variants with both properties have high potential to be causal and manifest effects on bodymass index through changes in chromatin structure. Causal determination will be made for a set of thesevariants using genome editing techniques such as CRISPR/Cas9 to generate isogenic human neuronal celllines that differ by genotype only at the single locus. Changes in exon-specific target gene expression andchromatin status will be assessed across the 3 developmental stages and under each obesogenic condition.Discovery of epigenetic mechanisms connected to genetic liability will translate the genetic risk information andidentify potential underlying factors behind both heritable and diet-induced obesity susceptibility.