Ca2+-dependent gene expression

In β-cells, Ca2+ signaling is critical for glucose-stimulated insulin secretion, but prolonged increases in Ca2+ due to chronic metabolic stress leads to β-cell dysfunction. By performing deep RNA sequencing on islets from several murine models of metabolic stress we have identified hundreds of genes whose expressions are adversely affected. Among these many genes we chose to study Ascl1, a basic helix-loop-helix transcription factor. The gene is critical for neurogenesis and is strongly increased during metabolic stress. Using a standard Cre/LoxP strategy, we recently discovered that the removal of Ascl1 from β-cells improves their function in response to metabolic stress by attenuating dedifferentiation, improving β-cell proliferation, and increasing parasympathetic innervation of islets. We anticipate continuing to study the maladaptive role of Ascl1 in islet function. In addition, to begin determining the function of some of the many other genes whose expression is perturbed by metabolic stress, we are implementing a high throughput method that relies both on CRISPR inhibition in mice and single cell RNA sequencing methods. We expect this strategy will provide insights into the function of over a dozen different Ca2+-dependent genes in the same experiment, accelerating our studies.