OR33-2 Unbiased Genome-wide Analyses Reveal Insulin Receptor Recruitment to Novel Genomic Sites in Pancreatic Islets

Program: Abstracts - Orals, Featured Poster Presentations, and Posters
Session: OR33-Insulin Signaling & Inflammation
Monday, June 17, 2013: 11:15 AM-12:45 PM
Presentation Start Time: 11:30 AM
Room 303 (Moscone Center)
Shweta Bhatt*1, Rachael Martinez2, Tze Howe Charn3, Michael Molla2 and Rohit N Kulkarni4
1Joslin Diabetes Center and Harvard Medical School, Boston, MA, 2Joslin Diabetes Center, Boston, MA, 3Stanford University, San Francisco, CA, 4Joslin Diabetes Center and Harvard Medical School, Boston, MA
Growth hormone receptors, like the Insulin Receptor (IR), are expressed ubiquitously and mediate critical metabolic actions of insulin in target tissues to regulate glucose homeostasis. Their precise role in the pathogenesis of metabolic disorders such as type 2 diabetes continues to be unraveled by the identification of novel downstream targets. We report, for the first time, the nuclear presence of IR protein in pancreatic beta cells and hepatocytes, two target tissues important for the regulation of whole body glucose homeostasis. Further investigation in pancreatic beta cells revealed a ligand (glucose and/or insulin, IGF1) and time dependent nuclear translocation of IR protein by live-cell imaging, immuno-staining and western blot analyses (n=3). Further, bioinformatics analyses revealed a consensus sequence (SRKRRS) in IR, conserved across species, and critical for its nuclear translocation, as confirmed by site-directed mutagenesis, nuclear-cytoplasmic fractionation and western blot analyses (n=3). Interestingly, this phenomenon was consistent in mouse, human islets and isolated beta cells and mutations associated with defects in insulin processing in humans. To explore the underlying molecular mechanism, we subjected mouse pancreatic islets to ex-vivo treatment with ligands (glucose and/or insulin, IGF1) followed by (a) genome-wide chromatin immuno-precipitation sequencing (ChIPseq) analysis for IR protein to assess genomic recruitment or (b) microarray gene expression analysis to study transcript regulation. Indeed, we observed IR recruitment to 9663 genomic sites, enriched in distal intergenic and intronic regions (similar to classic nuclear receptors), and associated with 6778 unique genes. Interestingly, a significant number of these genomic targets, several with established roles in glucose homeostasis (Mafa, Igf1r, Skp2, Insig2, Foxc1, Mapk1, Wnt5a, p21 etc.), were regulated by ligand in microarray and Q-PCR analyses, suggesting functional relevance (n=3; p<0.001). Consistently, this regulation was blocked in mouse islets or beta cells obtained from beta-cell specific IR knockout (bIRKO) animals and human islets subjected to shRNA mediated IR knockdown, suggesting an essential role of IR in this regulation. In summary, our unbiased approach identifies new primary targets of IR worthy of further investigation, to identify novel pathways and understand disease mechanism(s) for therapeutic intervention.

Nothing to Disclose: SB, RM, THC, MM, RNK

*Please take note of The Endocrine Society's News Embargo Policy at http://www.endo-society.org/endo2013/media.cfm

Sources of Research Support: NIH RO1 67536