Deacetylation of FoxO by Sirt1 Plays an Essential Role in Mediating Starvation-Induced Autophagy in Cardiac Myocytes

Abstract

Rationale: Autophagy, a bulk degradation process of cytosolic proteins and organelles, is protective during nutrient starvation in cardiomyocytes (CMs). However, the underlying signaling mechanism mediating autophagy is not well understood. Objective:We investigated the role of FoxOs and its posttranslational modification in mediating starvationinduced autophagy. Methods and Results:Glucose deprivation (GD) increased autophagic flux in cultured CMs, as evidenced by increased mRFP-GFP-LC3 puncta and decreases in p62, which was accompanied by upregulation of Sirt1 and FoxO1. Overexpression of either Sirt1 or FoxO1 was sufficient for inducing autophagic flux, whereas both Sirt1 and FoxO1 were required for GD-induced autophagy. GD increased deacetylation of FoxO1, and Sirt1 was required for GD-induced deacetylation of FoxO1. Overexpression of FoxO1(3A/LXXAA), which cannot interact with Sirt1, or p300, a histone acetylase, increased acetylation of FoxO1 and inhibited GD-induced autophagy. FoxO1 increased expression of Rab7, a small GTP-binding protein that mediates late autophagosome-lysosome fusion, which was both necessary and sufficient for mediating FoxO1-induced increases in autophagic flux. Although cardiac function was maintained in control mice after 48 hours of food starvation, it was significantly deteriorated in mice with cardiac-specific overexpression of FoxO1(3A/LXXAA), those with cardiac-specific homozygous deletion of FoxO1 (c-FoxO1 Key Words: autophagy Ⅲ starvation Ⅲ FoxO Ⅲ Sirt1 Ⅲ Rab7 Ⅲ deacetylation M acroautophagy (termed hereafter as autophagy) is a dynamic process of intracellular bulk degradation in which cytosolic proteins and organelles are sequestered into double-membrane vesicles called autophagosomes to be fused with lysosomes for degradation. 1 In the heart, autophagy maintains protein quality control, adapts to nutrient and oxygen deprivation during myocardial ischemia, and mediates cell death during reperfusion injury. 2,3 Autophagy during nutrient deprivation is an adaptive mechanism that allows the cells to survive by degrading the intracellular protein and lipid cargo and recycling the amino and fatty acids to generate ATP. 2 The nutrient status has a profound effect on cardiac contractility, and activation of autophagy during starvation is protective for the heart. The intracellular signaling mechanism by which nutrient starvation activates autophagy in cardiomyocytes (CMs) is not well understood, however.The forkhead box, class O (FoxO) family of transcription factors are present as 4 distinct isoforms (FoxO1, FoxO3, FoxO4, and FoxO6) in mammals. FoxO proteins play an important role in several intracellular functions, such as metabolism, stress resistance, longevity, tumor suppression, and cell size regulation. 4 The key to the myriad functions of FoxO proteins lies in the complex posttranslational modifications they undergo. They are phosphorylated in response to insulin and growth factors, dephosphorylated by protein phosphatases, ubiquitinated in response to oxidative stre...

Autophagy Induction by SIRT6 through Attenuation of Insulin-like Growth Factor Signaling Is Involved in the Regulation of Human Bronchial Epithelial Cell Senescence

“…Among the sirtuin family, both SIRT1 and SIRT6 are localized in the nucleus and share functional similarity in terms of antisenescence as relates to extending life span. Although SIRT1 induces autophagy for regulation of longevity in case of caloric restriction (30)(31)(32), reduced SIRT1 expression by CS exposure has been implicated in increased autophagy activation in COPD pathogenesis through enhanced apoptosis (11). Therefore, decreased expression levels of both SIRT1 and SIRT6 2) and SIRT6 cDNA (lanes 3, 4) transfected HBECs.…”

Metabolic effects of fasting on human and mouse blood in vivo

“…27,28 Therefore, we investigated the effects of starvation on protein acetylation levels by immunofluorescence staining of permeabilized and fixed leukocytes with an antibody specific for protein containing acetylated lysine residues. Cytofluorometric analyses revealed that starvation caused a significant reduction in the levels of protein acetylation in circulating white blood cells from mice ( Fig.…”

Repetitive stimulation of autophagy-lysosome machinery by intermittent fasting preconditions the myocardium to ischemia-reperfusion injury

“…Since fasting stimulates myocardial autophagy, [11][12][13]16 we evaluated cardiomyocyte autophagosome abundance in male mice carrying cardiomyocyte autophagy reporter GFP-tagged MAP1LC3B (green fluorescent protein-tagged microtubule-associated protein 1 light chain 3) transgene 13 subjected to intermittent (qOD) fasting for 6 wk (on a fed day). Mice subjected to 24 and 48 h of fasting, or to 24 h of fasting followed by 24 h of refeeding to mimic a fasting-refeeding cycle; and nonfasted agematched controls were studied in parallel.…”

“…27 We examined the effects of partial loss of LAMP2 on autophagy in the myocardium, which is a critical lysosomal pathway stimulated in response to starvation, 11,14,15 and is impaired in mice with complete loss of LAMP2. 28 Female Lamp2 heterozygous null mice (bearing the X/null genotype at the Lamp2 locus), demonstrate no difference in body weight or heart weight ( Table 1) as compared with littermate wild-type females.…”

“…total calorie restriction) is a potent stimulus for induction of myocardial macroautophagy (henceforth referred to as 'autophagy'). [11][12][13] Indeed, autophagy is essential for cardiac homeostasis in the perinatal starvation period at birth, prior to establishment of maternal milk supply; as mice with genetic ablation of autophagy proteins ATG5 and ATG7 cannot form autophagosomes and develop fatal myocardial ischemia. 14,15 Autophagy is also essential for maintenance of cardiac structure and function during prolonged starvation in mice, as concomitant impairment of autophagy with genetic ablation of Foxo1, haploinsufficiency of Becn1 11 or pharmacological inhibition with bafilomycin A 1 , 16 an inhibitor of lysosome acidification and function, results in rapid development of cardiomyopathy with starvation.…”