1728-P: Transcriptional Activation of Myc by Glucose Requires Recruitment of ChREBP to the Pvt1 Promoter Regulating Proliferation in ß-Cells

Myc and Carbohydrate Response Element (ChoRE) Binding Protein (ChREBP) are glucose-responsive transcriptional regulators of β-cell proliferation. Myc requires ChREBP to drive glucose-mediated proliferation in β-cells, yet no ChoREs are present in Myc’s promoter. In primates and rodents, the Myc gene and the long non-coding RNA, PVT1 gene are syntenic, separated by approximately 60 kbp. Mutations in this loci are associated with malignancies and complications of diabetes but are unexplored in β-cells. In INS-1 cells, Myc and PVT1 mRNAs were induced in response to glucose by 3.5±0.02and 2.9±0.1 fold, respectively, and Myc protein levels increased by 4-fold ±0.17. Silencing of ChREBPβ significantly blocked the glucose response of Myc and Pvt1. ChREBP ChIPseq revealed a single ChoRE located approximately 100 bp upstream of the Pvt1 transcription start site and is highly conserved in primates and rodents. By Chromosome Conformation Capture (3C), we demonstrated that high glucose increased interaction between the Pvt1 and Myc promoters by a factor of 3.5±0.4. Using adenoviral delivery of CRISPR-xCas9-sgRNA targeting the Pvt1 ChoRE, we specifically attenuated ChREBP recruitment by 2.8±0.8 fold, while other known ChoREs were unaffected. We demonstrated by 3C that the chromatin looping in response to glucose was abolished by the Pvt1 ChoRE disruptor. Both in primary rat islets and in INS-1 cells, disruption of Pvt1’s ChoRE resulted in complete and significant blockage of Pvt1 and Myc mRNA upregulation in response to glucose, with no effect on other ChoRE-containing genes. Finally, disruption of the ChoRE on the Pvt1 promoter eliminated glucose-mediated proliferation of primary rat β-cells (a decrease of 9.8±1.9 fold on Ki67+ Ins+ staining). In conclusion, we identify a novel mechanism driving the ChREBP-dependent activation of the Myc and Pvt1 genes and proliferation of β-cells. Disclosure L.S.Katz: None. P.Wang: None. G.Brill: None. P.Zhang: None. J.Haldeman: Employee; Janssen Research & Development, LLC. C.B.Newgard: Advisory Panel; Eli Lilly and Company, Novo Nordisk, AstraZeneca. A.F.Stewart: None. A.Garcia-ocana: Consultant; Sun Pharmaceutical Industries Ltd. D.Scott: None. Funding National Institutes of Health (R01DK130300, P30DK020541)