Autophagy: Intro to the concepts.
Autophagy is divided into three theoretical frame works, macroautophagy, microautophagy, and chaperone mediated autophagy. Autophagocytosis of the ER is thought to occur through all of these methods. Macroautophagy comes in two flavors, the first being a nonspecific autophagy that occurs in response to starvation where cytoplasm is indiscriminately internalized into autophagosomes so that it can be recycled to feed the cell. Selective macroautophagy involves autophagosomal internalization of organelles and cellular components that have been marked for destruction and occurs even during nutrient rich conditions. Unlike macroautophagy, microautophagy and chaperone mediated autophagy do not involve the autophagosome. Instead, both occur by direct engulfment of cellular material by the lysosome. Microautophagy is a random sampling of cellular cytoplasm by the lysosome, whereas chaperone mediated autophagocytosis involves a more complex signaling network centering around the hsc70 complex with binds targets to the lysosomal network\cite{Agarraberes1997}. This review will discuss selective macroautophagy in the greatest depth as this field has seen many interesting advances, with respect to the ER, in recent years. We will also discuss some micro and chaperone mediated autophagocytosis.
Selective autophagy is akin to cellular housekeeping where organelles and other cellular components that have become damaged, toxic, or are no longer required by the cell are recycled. A diverse set of organelles and cellular components have been shown to be recycled by selective autophagocytosis ranging from old mitochondria to toxic protein aggregates. For each recycled component there are autophagy receptors which bind to specific cargo linking it to the autophagosomal machinery\cite{Zaffagnini2016}. The autophagosomal machinery (proteins involved) in turn helps recruit the isolation membrane and form the autophagophore. This membrane fuses with itself, internalizing the marked cargo and forming the double membrane autophagosome. The autophagosome then fuses with the lysosome in the case of mammals or the vacuole in the case of yeast, delivering the cargo where upon it is broken down into its molecular constituents.
ERphagy In Yeast:
ERphagy and ERAD:
Selective ERphagy is tightly linked to ER stress and in particular the unfolded protein response (UPR)\cite{Yorimitsu2006,Kruse2006,Kamimoto2006,Kouroku2007}. ER stress causes a significant ER expansion, and is also associated with an increase in autophagosome numbers. While this connection was made in the early 1970’s only recently has the mechanism behind these observations been studied. UPR is initiated by an overabundance of misfolded proteins in the ER lumen. This occurs when cellular demand for membrane/secreted proteins exceeds the ER’s ability to facilitate protein folding. Bernales et al. asked what changes in ER morphology happened upon UPR induction in yeast by dithiothreitol\cite{Bernales2007}. Using thin section electron microcopy, they observed a large expansion of ER volume and a change in morphology biasing the ER towards large sheet domains. Upon further examination, Bernales et al. noted that in a subset of cells UPR resulted in a marked increase in double membrane autophagosome like structures packed with membrane cisternae. They hypothesized that these were autophagosomes which had specifically internalized ER membrane in response to UPR. Bernales et al. went on to show that these structures did indeed contain ER membrane and that the formation of these structures was dependent on autophagosomal machinery. When the autophagosomal machinery was knocked out the cells were no longer able to tolerate dithiothreitol induced UPR suggesting that ERphagy is an essential component of UPR. Interestingly, the degradation of the membrane contents seemed to be unnecessary for the survival of cells. As long as the ER membrane was packed in autophagosomes the cells were able to tolerate the dithiothreitol induced UPR.
ERphagy and ER Stress:
ERphagy has also been connected with the clearance of large protein aggregates in the ER. Kruse et al. asked what cellular processes were implicated in a liver disease associated with a mutant aggregate prone (AP) fibrinogen\cite{Kruse2006}. They showed that upon high expression of AP fibrinogen proteasomal degradation was no longer sufficient to clear aggregates. In these high expression conditions ERphagy was required to clear the AP fibrinogen. This implicates ERphagy not only in the liver disease but also basic protein quality control and ER homeostasis.
Further investigation of the role of ERphagy in maintaining ER homeostasis by Schuck et al. highlighted a role for ERphagy in basic ER stress response\cite{Schuck2014}. They showed that, tunicamycin induced ER stress is associated with an increase in not only ER size but also abnormal ER structures they term ER whorls. They observed by electron microscopy that ER whorls were internalized directly by the vacuole. Through knockout they discovered that this form of microautophagy of the ER is not dependent on any of the canonical autophagy genes (atg1, atg6, atg7, atg8, atg14, atg16). Linking Pho8D60, a marker of vacuole internalization, to ER and cytoplasmic proteins Schuck et al. were able to demonstrate that ER stress results in the selective increase in ERphagy and not a general increase in autophagy. This indicates that ER stress was able to activate an alternative ER specific microautophagy and not other forms of generally autophagy. The signaling cascade involved in the recognition of ER whorls, their separation from the rest of the ER membrane, binding to the vacuole, and internalization has not yet been elucidated.