Mapping of long-range INS promoter interactions reveals a role for calcium-activated chloride channel ANO1 in insulin secretion

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 111; no. 47; pp. 16760 - 16765
• Main Authors: Xu, Zhixiong, Lefevre, Gaelle M., Gavrilova, Oksana, St. Claire, Mark B. Foster, Riddick, Gregory, Felsenfeld, Gary
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 25.11.2014; National Acad Sciences
• Subjects: Animals; Anoctamin-1; Biological Sciences; Calcium; Chloride channels; Chloride Channels - genetics; Chloride Channels - physiology; Chlorides; Chromatin; Chromosomes; Gene expression; Genes; Genetic loci; Genomics; Glucose - metabolism; Humans; Insulin; Insulin - genetics; Insulin - metabolism; Insulin Secretion; Islets of Langerhans; Islets of Langerhans - metabolism; Mice; Mice, Knockout; Neoplasm Proteins - genetics; Neoplasm Proteins - physiology; Polymerase Chain Reaction; Promoter Regions, Genetic; Rodents; Small interfering RNA; Social interaction; chloride channel; diabetes; insulin secretion
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1419240111

Significance Within the cell nucleus, the genetic material is organized into loop domains that transiently bring together genes separated by various distances along linear DNA. We asked whether the insulin ( INS ) gene in human pancreatic beta cells, which secrete the insulin protein, makes contact with other genes that play some role in insulin function. We show that the INS promoter contacts the anoctamin 1 (ANO1) gene, located far away on the same chromosome, stimulating its expression, and that ANO1, a chloride ion channel protein, plays a role in insulin secretion. Measurements of long-range interactions detect regulatory pathways not evident from other kinds of studies. In this case, such experiments reveal previously unrecognized mechanisms that could affect susceptibility to human diabetes. We used circular chromatin conformation capture (4C) to identify a physical contact in human pancreatic islets between the region near the insulin ( INS ) promoter and the ANO1 gene, lying 68 Mb away on human chromosome 11, which encodes a Ca ²⁺-dependent chloride ion channel. In response to glucose, this contact was strengthened and ANO1 expression increased, whereas inhibition of INS gene transcription by INS promoter targeting siRNA decreased ANO1 expression, revealing a regulatory effect of INS promoter on ANO1 expression. Knockdown of ANO1 expression caused decreased insulin secretion in human islets, establishing a physical proximity-dependent feedback loop involving INS transcription, ANO1 expression, and insulin secretion. To explore a possible role of ANO1 in insulin metabolism, we carried out experiments in Ano1 ⁺/⁻ mice. We observed reduced serum insulin levels and insulin-to-glucose ratios in high-fat diet–fed Ano1 ⁺/⁻ mice relative to Ano1 ⁺/⁺ mice fed the same diet. Our results show that determination of long-range contacts within the nucleus can be used to detect novel and physiologically relevant mechanisms. They also show that networks of long-range physical contacts are important to the regulation of insulin metabolism.