Search

Recent studies have uncovered thousands of long non-coding RNAs (lncRNAs) in human pancreatic beta cells. beta cell lncRNAs are often cell type specific and exhibit dynamic regulation during differentiation or upon changing glucose concentrations. Although these features hint at a role of lncRNAs in beta cell gene regulation and diabetes, the function of beta cell lncRNAs remains largely unknown. In this study, we investigated the function of beta cell-specific lncRNAs and transcription factors using transcript knockdowns and co-expression network analysis. This revealed lncRNAs that function in concert with transcription factors to regulate b cell-specific transcriptional networks. We further demonstrate that the lncRNA PLUTO affects local 3D chromatin structure and transcription of PDX1, encoding a key b cell transcription factor, and that both PLUTO and PDX1 are downregulated in islets from donors with type 2 diabetes or impaired glucose tolerance. These results implicate lncRNAs in the regulation of beta cell-specific transcription factor networks.

International audience; Reserpine treatment leads to a trans-synaptic increase of the tyrosine hydroxylase (TH) gene transcription rate, mRNA and protein levels in catecholaminergic tissues including the adrenal medulla (AM) and the superior cervical ganglia (SCG). The TPA-responsive element plays an important role in the trans-synaptically-induced transcription of the TH gene in the AM, whereas it does not appear to be involved in the SCG (Trocmé et al. [1997] J. Neurosci. Res. 48:489-498). In this study, we show that another regulatory sequence of the TH proximal promoter, the cAMP-responsive element (CRE), binds different factors in the AM and in the SCG. To elucidate the dynamics of promoter regulation a complete time course analysis was conducted. Reserpine treatment enhances, between 1 hr and 8 hr after the injection, the expression and the binding of the repressor ICER in the AM, whereas in the SCG it enhances the binding of CREM factors. These results suggest that the mechanisms mediating trans-synaptic induction of the TH gene are different in the AM and SCG. The interplay between positive and negative transcription factors and their kinetics of action are responsive of the long-term regulation of the TH gene.

Summary Despite the central role of chromosomal context in gene transcription, human noncoding DNA variants are generally studied outside of their genomic location. This limits our understanding of disease-causing regulatory variants. INS promoter mutations cause recessive neonatal diabetes. We show that all INS promoter point mutations in 60 patients disrupt a CC dinucleotide, whereas none affect other elements important for episomal promoter function. To model CC mutations, we humanized an ∼3.1-kb region of the mouse Ins2 gene. This recapitulated developmental chromatin states and cell-specific transcription. A CC mutant allele, however, abrogated active chromatin formation during pancreas development. A search for transcription factors acting through this element revealed that another neonatal diabetes gene product, GLIS3, has a pioneer-like ability to derepress INS chromatin, which is hampered by the CC mutation. Our in vivo analysis, therefore, connects two human genetic defects in an essential mechanism for developmental activation of the INS gene., Graphical abstract, Highlights • Mutations of a CC dinucleotide in the human INS promoter cause neonatal diabetes • We humanized ∼3.1 kb of mouse Ins2 and created a CC mutant version • Humanized Ins2, but not the CC mutant, recapitulates developmental chromatin activation • GLIS3, also mutated in diabetes, activates INS chromatin and requires an intact CC, Mutations in the CC element of the INS promoter or the transcription factor GLIS3 cause neonatal diabetes. Akerman et al. humanize a 3.1-kb region upstream of the mouse Ins2 gene and show that GLIS3 and the CC element form a pioneering mechanism that activates INS chromatin during pancreas development.

Eph/ephrin interactions and their bidirectional signaling are integral part of the complex communication system between β-cells, essential for glucose homeostasis. Indeed, Eph/ephrin system was shown to be directly involved in the glucose-stimulated insulin secretion (GSIS) process occurring in the pancreatic islets. Here we tested the Eph antagonist UniPR500 as GSIS enhancer. UniPR500 was validated as EphA5-ephrin-A5 inhibitor in vitro and its efficacy as GSIS enhancer was assessed on EndoC-βH1 cells. The selectivity of UniPR500 was evaluated by testing this compound on a panel of well-known molecular targets responsible for the regulation of glucose homeostasis. Plasmatic levels of UniPR500 were measured by HPLC/MS approach after oral administration. Finally, UniPR500 was tested as hypoglycemic agent in healthy mice, in a non-genetic mouse model of insulin resistance (IR) and in a non-genetic mouse model of type 1 diabetes (T1D). The compound is an orally bioavailable and selective Eph antagonist, able to increase GSIS from EndoC-βH1 cells. When tested in vivo UniPR500 showed to improve glucose tolerance in healthy and IR mice. As expected by a GSIS enhancer acting on healthy β-cells, UniPR500 was ineffective when tested on a non-genetic mouse model of type 1 diabetes, where pancreatic function was severely compromised. In conclusion our findings suggest that Eph targeting is a new and valuable pharmacological strategy in the search of new hypoglycemic agents.

Summary Recent studies have uncovered thousands of long non-coding RNAs (lncRNAs) in human pancreatic β cells. β cell lncRNAs are often cell type specific and exhibit dynamic regulation during differentiation or upon changing glucose concentrations. Although these features hint at a role of lncRNAs in β cell gene regulation and diabetes, the function of β cell lncRNAs remains largely unknown. In this study, we investigated the function of β cell-specific lncRNAs and transcription factors using transcript knockdowns and co-expression network analysis. This revealed lncRNAs that function in concert with transcription factors to regulate β cell-specific transcriptional networks. We further demonstrate that the lncRNA PLUTO affects local 3D chromatin structure and transcription of PDX1, encoding a key β cell transcription factor, and that both PLUTO and PDX1 are downregulated in islets from donors with type 2 diabetes or impaired glucose tolerance. These results implicate lncRNAs in the regulation of β cell-specific transcription factor networks., Graphical Abstract, Highlights • A loss-of-function screen reveals functional β cell lncRNAs • Cell-specific lncRNAs and transcription factors regulate common gene networks • The lncRNA PLUTO influences interactions between an enhancer cluster and PDX1 • PLUTO and PDX1 are deregulated in type 2 diabetes and impaired glucose tolerance, Akerman et al. studied the function of human β cell lncRNAs with RNAi, CRISPRi, and co-expression networks. This revealed β cell lncRNAs and transcription factors that control common regulatory networks. One lncRNA, PLUTO, is downregulated in type 2 diabetes and controls PDX1, encoding a key β cell transcription factor.