Oxidative protein folding in the endoplasmic reticulum: tight links to the mitochondria-associated membrane (MAM)

Abstract

The production of secretory proteins at the ER (endoplasmic reticulum) depends on a ready supply of energy and metabolites as well as the close monitoring of the chemical conditions that favor oxidative protein folding. ER oxidoreductases and chaperones fold nascent proteins into their export-competent three-dimensional structure. Interference with these protein folding enzymes leads to the accumulation of unfolded proteins within the ER lumen, causing an acute organellar stress that triggers the UPR (unfolded protein response). The UPR increases the transcription of ER chaperones commensurate with the load of newly synthesized proteins and can protect the cell from ER stress. Persistant stress, however, can force the UPR to commit cells to undergo apoptotic cell death, which requires the emptying of ER calcium stores. Conversely, a continuous ebb and flow of calcium occurs between the ER and mitochondria during resting conditions on a domain of the ER that forms close contacts with mitochondria, the MAM (mitochondria-associated membrane). On the MAM, ER folding chaperones such as calnexin and calreticulin and oxidoreductases such as ERp44, ERp57 and Ero1alpha regulate calcium flux from the ER through reversible, calcium and redox-dependent interactions with IP3Rs (inositol 1,4,5-trisphophate receptors) and with SERCAs (sarcoplasmic/endoplasmic reticulum calcium ATPases). During apoptosis progression and depending on the identity of the ER chaperone and oxidoreductase, these interactions increase or decrease, suggesting that the extent of MAM targeting of ER chaperones and oxidoreductases could shift the readout of ER-mitochondria calcium exchange from housekeeping to apoptotic. However, little is known about the cytosolic factors that mediate the on/off interactions between ER chaperones and oxidoreductases with ER calcium channels and pumps. One candidate regulator is the multi-functional molecule PACS-2 (phosphofurin acidic cluster sorting protein-2). Recent studies suggest that PACS-2 mediates localization of a mobile pool of calnexin to the MAM in addition to regulating homeostatic ER calcium signaling as well as MAM integrity. Together, these findings suggest that cytosolic, membrane and lumenal proteins combine to form a two-way switch that determines the rate of protein secretion by providing ions and metabolites and that appears to participate in the pro-apoptotic ER-mitochondria calcium transfer.

Figure 1

Three major metabolic exchanges between the ER and mitochondria impact on ER chaperone systems that catalyze the production of secretory proteins. 1. Energy exchange. GRP78/BiP requires ATP that is predominantly supplied by mitochondria, and to a lesser extent from glycolysis. GRP78/BiP also depends on ER calcium. 2. Oxidative protein folding. PDI forms disulfide bonds with the help of its recharger Ero1α, which can bind calcium and FAD derived from mitochondrial metabolism. Import proteins of FAD into the ER are currently unknown in mammalian systems. ROS produced by Ero1α could directly impact on the mitochondrial permeability transition pore (MPTP), SERCA and the IP3R. 3. Calcium flux. The calnexin/calreticulin folding cycle depends on ATP and ER calcium. Calnexin and calreticulin buffer free calcium within the ER, but also associate with SERCA to modulate calcium import. The localization of calnexin to the MAM and hence potentially its interaction with SERCA is under the control of PACS-2. The MAM-associated moiety of the oxidoreductase ERp44 interacts with the IP3R and regulates IP3R calcium release.