Molecular mechanisms of autophagy in the cardiovascular system

Abstract

Autophagy is a catabolic recycling pathway triggered by various intra- or extracellular stimuli that is conserved from yeast to mammals. During autophagy, diverse cytosolic constituents are enveloped by double-membrane vesicles, autophagosomes, which later fuse with lysosomes or the vacuole to degrade their cargo. Dysregulation in autophagy is associated with a diverse range of pathologies including cardiovascular disease, the leading cause of death in the world. As such, there is great interest in identifying novel mechanisms that govern the cardiovascular response to disease-related stress. First described in failing hearts, autophagy within the cardiovascular system has been characterized widely in cardiomyocytes, cardiac fibroblasts, endothelial cells, and vascular smooth muscle cells. In all cases, a window of optimal autophagic activity seems to be critical to the maintenance of cardiovascular homeostasis and function; excessive or insufficient levels of autophagic flux can each contribute to heart disease pathogenesis. Here, we review the molecular mechanisms that govern autophagosome formation and analyze the link between autophagy and cardiovascular disease.

Keywords: atherosclerosis; blood vessels; cardiovascular diseases; heart; intracellular signaling proteins; muscle, smooth, vascular; myocytes, cardiac.

Figure 1. The three main types of autophagy

(A) Yeast cells carry out both macroautophagy and microautophagy. While macroautophagy consists of the bulk degradation of cytoplasmic material that is sequestered inside double-membrane autophagosomes that then fuse with the vacuole, microautophagy works by directly taking up the substrates through invagination of the vacuole. (B) In mammals along with microautophagy and macroautophagy, chaperone-mediated autophagy enables the degradation of specific protein substrates that contain a KFERQ motif that is recognized by chaperones that mediate the translocation of the protein into the lysosome through LAMP2A.

Figure 1. The three main types of autophagy

(A) Yeast cells carry out both macroautophagy and microautophagy. While macroautophagy consists of the bulk degradation of cytoplasmic material that is sequestered inside double-membrane autophagosomes that then fuse with the vacuole, microautophagy works by directly taking up the substrates through invagination of the vacuole. (B) In mammals along with microautophagy and macroautophagy, chaperone-mediated autophagy enables the degradation of specific protein substrates that contain a KFERQ motif that is recognized by chaperones that mediate the translocation of the protein into the lysosome through LAMP2A.

Figure 2. Autophagy induction

(A) In yeast in nutrient-rich conditions TORC1 and PKA inhibit autophagy by phosphorylating Atg1 and Atg13. During starvation the Atg1 kinase complex is no longer repressed, Atg13 is partially dephosphorylated and Atg1 is activated. Atg1 then phosphorylates itself and other targets to induce autophagy. (B) In mammals in nutrient-rich conditions MTORC1 directly binds ULK1 through RPTOR and inhibits ULK1/2 and ATG13 by phosphorylation. Upon starvation MTORC1 dissociates from the ULK1 kinase complex, allowing ATG13 dephosphorylation and activating ULK1/2 that then phosphorylates members of the complex and other targets to induce autophagy.

Figure 2. Autophagy induction

(A) In yeast in nutrient-rich conditions TORC1 and PKA inhibit autophagy by phosphorylating Atg1 and Atg13. During starvation the Atg1 kinase complex is no longer repressed, Atg13 is partially dephosphorylated and Atg1 is activated. Atg1 then phosphorylates itself and other targets to induce autophagy. (B) In mammals in nutrient-rich conditions MTORC1 directly binds ULK1 through RPTOR and inhibits ULK1/2 and ATG13 by phosphorylation. Upon starvation MTORC1 dissociates from the ULK1 kinase complex, allowing ATG13 dephosphorylation and activating ULK1/2 that then phosphorylates members of the complex and other targets to induce autophagy.

Figure 3. Autophagy regulation

Through STK11/LKB1, AMPK senses decreases in the ATP/AMP ratio and phosphorylates TSC1-TSC2, which then targets RHEB, leading to MTORC1 inhibition and autophagy activation. INSR/IGF1R triggers the activation of the class I PI3K, inducing the formation of phosphatidylinositol(3,4,5)triphosphate (PIP₃) and AKT/PKB activation; AKT can inhibit TSC1/TSC2, blocking autophagy. PTEN works as a PIP₃ phosphatase generating phosphatidylinositol(4,5)bisphosphate (PIP₂) and inducing autophagy.

Figure 4. Class III PtdIns3K complexes

Three class III PtdIns3K complexes can be observed in mammals. All of them require PIK3C3/VPS34, PIK3R4/VPS15 and BECN1. Specific subunits regulate the function of the different complexes. Binding of ATG14 and AMBRA1 leads to autophagy induction. UVRAG and SH3GLB1 binding also activates autophagy, whereas binding to KIAA0226/RUBICON inhibits autophagosome maturation.

Figure 5. Autophagosomes have a diverse range of potential membrane sources

The trans-Golgi Network, mitochondria, mitochondrial associated membrane and ER have been postulated as membrane donors. Omegasomes have been described as the ER structures that work as a platform for autophagosome formation. The phagophore (shown in red) elongates and engulfs part of a cisternae before it buds off the ER and becomes an autophagosome.

Figure 6. Two ubiquitin-like conjugation systems

Atg8 and Atg12 go through subsequent activation, mediated by Atg7 and conjugation mediated by Atg10 and Atg3, respectively, before covalently binding to PE in the case of Atg8, and Atg5 in the case of Atg12. Atg8–PE binds both the inner and outer membrane of the autophagosome, but can be deconjugated by Atg4, the same protein that removes the C-terminal arginine initially present at the Atg8 C terminus. Atg12–Atg5 bind Atg16 creating a large multimeric complex that locates to the phagophore and enhances Atg8 lipidation and membrane expansion.

Figure 7. Autophagy regulators in cardiomyocytes

A wide variety of stimuli can regulate autophagy in cardiomyocytes. Some of them are autophagy activators and are associated with cardiovascular diseases. However, acetylcholine, catecholamines, aging, IGF1 and INS/insulin are capable of inhibiting autophagy. INS and IGF1 are well known cardioprotective agents. AchR, acetylcholine receptor; AGT II, angiotensin II.

Figure 8. Autophagy in cardiovascular diseases

The relationship between autophagy and cardiovascular diseases is complex. Although basal autophagy is critical to maintain cell homeostasis, both increases and decreases in autophagy to an excessive degree can induce alterations in normal heart and blood vessel functions. In ischemia/reperfusion, heart failure, hypertrophy, diabetes, atherosclerosis, plaque destabilization, lesional thrombosis and vascular smooth muscle cell (VSMC) proliferation, autophagic flux is abnormally elevated, contributing to cardiac and vessel dysfunction. In Atg5 and Becn1 knockout animals and during aging, autophagic activity is decreased, perturbing cellular homeostasis and contributing to cardiovascular diseases, such as post-operative atrial fibrillation (POAF), ischemia-induced damage, hypertrophy, heart failure, and vascular endothelial cell dysfunction.