Identification of Insulin-Responsive Transcription Factors Regulating Hepatocyte Glucose Production

Hepatocyte glucose production is a complex process that integrates cell-autonomous mechanisms with cellular signaling, modulation of enzyme activities, and gene transcription. Transcriptional mechanisms controlling glucose production are redundant, and involve nuclear hormone receptors as well as unliganded transcription factors (TFs). Our knowledge of this circuitry is incomplete. Here we used DNA affinity purification followed by mass spectrometry to probe the network of hormone-regulated TFs using Phosphoenolpyruvate carboxykinase ( Pck1 ) and Glucose-6-phosphatase ( G6pc ) in liver as well as primary hepatocytes as model systems. The repertoire of insulin-regulated TFs is unexpectedly broad and diverse. Whereas in liver the two test promoters are regulated by largely overlapping sets of TFs, in primary hepatocytes Pck1 and G6pc regulation diverge. Insulin treatment preferentially results in increased occupancy of the two promoters, consistent with a model in which the hormone’s primary role is to recruit co-repressors, rather than to clear activators. Nine insulin-responsive TFs are present in both models, but only FoxK1, FoxA2, ZFP91, and ZHX3 require an intact Pck1p insulin response sequence for binding. Knockdown of FoxK1 in primary hepatocytes decreased glucose production as well as insulin’s ability to suppress it. The findings expand the repertoire of insulin-dependent TFs, and identify FoxK1 as a contributor to insulin signaling.