FIGURE 8.

Increased binding of FoxO1 and FoxO3 to autophagy gene regulatory sequences in starved cardiomyocytes. Chromatin immunoprecipitation was performed on fed and starved cardiomyocytes. Cross-linked DNA-protein complexes from fed and starved cardiomyocytes were immunoprecipitated with either FoxO1, FoxO3, or IgG (negative control) antibodies. Precipitated DNA was then PCR-amplified using primers specific for Gabarpl1 and Atg12 gene regulatory sequences containing identified FoxO-binding sites. Increased binding of FoxO3 to Atg12 (2.7-fold) and Gabarapl1 (1.7-fold) promoters was observed under starvation conditions relative to fed controls. Increased FoxO1 binding was observed in both conditions (1.5-fold for Gabarapl1 and 1.65-fold for Atg12), but induction was reduced relative to FoxO3 compared with fed cardiomyocytes. Input represents cross-linked DNA amplified without immunoprecipitation with respective antibodies.