Autophagosomes form at ER–mitochondria contact sites

Abstract

Autophagy is a tightly regulated intracellular bulk degradation/recycling system that has fundamental roles in cellular homeostasis. Autophagy is initiated by isolation membranes, which form and elongate as they engulf portions of the cytoplasm and organelles. Eventually isolation membranes close to form double membrane-bound autophagosomes and fuse with lysosomes to degrade their contents. The physiological role of autophagy has been determined since its discovery, but the origin of autophagosomal membranes has remained unclear. At present, there is much controversy about the organelle from which the membranes originate--the endoplasmic reticulum (ER), mitochondria and plasma membrane. Here we show that autophagosomes form at the ER-mitochondria contact site in mammalian cells. Imaging data reveal that the pre-autophagosome/autophagosome marker ATG14 (also known as ATG14L) relocalizes to the ER-mitochondria contact site after starvation, and the autophagosome-formation marker ATG5 also localizes at the site until formation is complete. Subcellular fractionation showed that ATG14 co-fractionates in the mitochondria-associated ER membrane fraction under starvation conditions. Disruption of the ER-mitochondria contact site prevents the formation of ATG14 puncta. The ER-resident SNARE protein syntaxin 17 (STX17) binds ATG14 and recruits it to the ER-mitochondria contact site. These results provide new insight into organelle biogenesis by demonstrating that the ER-mitochondria contact site is important in autophagosome formation.

A current perspective of autophagosome biogenesis

“…Consistent with Atg14L dot formation on the ER, it was found that ULK1 and Atg5 also form dot-like accumulations on the ER, and that these dots are also associated with the ER when visualized by live cell imaging [20,68]. Recently, Koyama-Honda et al [69] have shown that ULK1 and Atg5 are simultaneously recruited and accumulate to form dots upon starvation, possibly through the direct interaction between FIP200 and Atg16L1 [46][47][48].…”

“…These two separate models, however, may be reconciled by the recent work by Hamasaki et al [68], which focuses on the contact sites between the ER and mitochondria (Figure 2). ER-mitochondria contact sites were previously known to exist and play important roles in mitochondria fission, calcium signaling, lipid transfer and so on [74].…”

Loss of sirtuin 1 and mitofusin 2 contributes to enhanced ischemia/reperfusion injury in aged livers

“…One potential mechanism behind MFN2‐induced autophagy could be linked to its unique localization at the mitochondria‐ER contact site, which is considered as a source for autophagosomal membranes during elongation (Hailey et al., 2010). Absence of mitochondria‐ER interface proteins such as MFN2 and phosphofurin acidic cluster sorting protein 2 (PACS2) prevents proper autophagosome formation in response to autophagic stimuli including starvation (Hamasaki et al., 2013) or ER stress (Muñoz et al., 2013). Moreover, in a recent study, the ER acetylation machinery directly regulates autophagy in the brain (Peng et al., 2016).…”

Autophagy at the crossroads of catabolism and anabolism

“…Although evidence supports the idea that nucleation of the isolation membrane occurs at a distinct site emanating from the endoplasmic reticulum (ER), termed the omegasome 17 , other sources of membrane contribute to autophagosome formation, including ER-Golgi intermediate compartments, ERmitochondri a junctions, mitochondria, endosomes and the plasma membrane [17][18][19][20][21][22] . Taken together, these studies highlight the complexities of autophagy initiation in mammals.…”