PROJECT SUMMARY Diabetic cardiomyopathy is the leading cause of complications in type-2 diabetes. The diabetic heart features metabolic insults that promote its metabolic inflexibility and failure, particularly insulin resistance, impaired glu- cose oxidation and loss of NAD+ biogenesis. The cardiomyocyte-specific glucocorticoid receptor (GR) is im- portant for heart function, but the GR-dependent metabolic mechanisms of glucocorticoids (GCs) in cardiomyo- cytes remain unknown. GR activity is regulated by the circadian clock but the extent to which the GR regulates the cardiomyocyte clock is still unknown. The GR activates its co-factor KLF15, which regulates cardiac metab- olism and promotes oscillating transcriptional programs in heart. However, a direct role for KLF15 in regulating circadian clock genes is still unknown. The circadian clock factor BMAL1 promotes oscillating NAD+ repletion and supports cardiac clock and function in a cardiomyocyte-autonomous fashion. However, the clock response to GC signaling in heart remains unknown. Recently, we have shown two interconnected time dimensions that harness benefits from detriments of GC pharmacology: circadian time-of-intake and chronic frequency-of- intake. Circadian-specific prednisone intermittence has shown positive outcomes of safety and benefits in a pilot clinical trial, underscoring relevance and feasibility in humans. Our preliminary findings with obese diabetic mice suggest that light-phase intermittent prednisone rescues NAD+ biogenesis, glucose metabolism and ceramide lipotoxicity in heart, blunting diastolic dysfunction. This raises the unanticipated idea of repurposing GC drugs to treat diabetic cardiomyopathy, but the mechanism underpinning this action must still be identified. Based on our findings in three cardiomyocyte-restricted inducible KO models, we discovered a new GR-KLF15-BMAL1 mechanism regulating clock and metabolism in cardiomyocytes. We test here the overarching hypothe- sis that this cardiomyocyte-autonomous axis is triggered by specific GC timing and rescues cardiomy- opathy in type-2 diabetes. In Aim 1, we will identify the circadian cue gating the circadian-specific effect of exogenous glucocorticoids in heart. We will test the hypothesis that the rhythmic trough of endogenous GCs is the circadian cue mediating the circadian-specific effects of exogenous GCs in heart. In Aim 2, we will determine the extent to which the cardiomyocyte clock discriminates beneficial versus deleterious outcomes of chronic GC frequency. We will test the hypothesis that cardiomyocyte-specific BMAL1 is the discriminating factor in the dia- betic heart response to chronic GC effects. In Aim 3, we will identify the mechanisms promoting insulin sensitivity and glucose oxidation by the cardiomyocyte-specific GR-KLF15 axis. We will test the hypothesis that the cardi- omyocyte GR-KLF15 axis rescues metabolic flexibility in the diabetic heart through a concerted program of ceramide reduction ...