Numerous nascent proteins undergo folding and maturation within the luminal and membrane compartments of the endoplasmic reticulum (ER). Despite the presence of various factors in the ER that promote protein folding, many proteins fail to properly fold and assemble and are subsequently degraded. Regulatory proteins in the ER also undergo degradation in a way that is responsive to stimuli or the changing needs of the cell. As in most cellular compartments, the ubiquitin-proteasome system (UPS) is responsible for the majority of the degradation at the ER-in a process termed ER-associated degradation (ERAD). Autophagic processes utilizing ubiquitin-like protein-conjugating systems also play roles in protein degradation at the ER. The ER is continuous with the nuclear envelope (NE), which consists of the outer nuclear membrane (ONM) and inner nuclear membrane (INM). While ERAD is known also to occur at the NE, only some of the ERAD ubiquitin-ligation pathways function at the INM. Protein degradation machineries in the ER/NE target a wide variety of substrates in multiple cellular compartments, including the cytoplasm, nucleoplasm, ER lumen, ER membrane, and the NE. Here, we review the protein degradation machineries of the ER and NE and the underlying mechanisms dictating recognition and processing of substrates by these machineries.

The E3 ligase membrane-associated ring-CH-type finger 6 (MARCH6) is a polytopic enzyme bound to the membranes of the endoplasmic reticulum. It controls levels of several known protein substrates, including a key enzyme in cholesterol synthesis, squalene monooxygenase. However, beyond its own autodegradation, little is known about how MARCH6 itself is regulated. Using CRISPR/Cas9 gene-editing, MARCH6 overexpression, and immunoblotting, we found here that cholesterol stabilizes MARCH6 protein endogenously and in HEK293 cells that stably express MARCH6. Conversely, MARCH6-deficient HEK293 and HeLa cells lost their ability to degrade squalene monooxygenase in a cholesterol-dependent manner. The ability of cholesterol to boost MARCH6 did not seem to involve a putative sterol-sensing domain in this E3 ligase, but was abolished when either membrane extraction by valosin-containing protein (VCP/p97) or proteasomal degradation was inhibited. Furthermore, cholesterol-mediated stabilization was absent in two MARCH6 mutants that are unable to degrade themselves, indicating that cholesterol stabilizes MARCH6 protein by preventing its autodegradation. Experiments with chemical chaperones suggested that this likely occurs through a conformational change in MARCH6 upon cholesterol addition. Moreover, cholesterol reduced the levels of at least three known MARCH6 substrates, indicating that cholesterol-mediated MARCH6 stabilization increases its activity. Our findings highlight an important new role for cholesterol in controlling levels of proteins, extending the known repertoire of cholesterol homeostasis players.

Endoplasmic reticulum (ER) stress occurs when the abundance of unfolded proteins in the ER exceeds the capacity of the folding machinery. Despite the expanding cadre of characterized cellular adaptations to ER stress, knowledge of the effects of ER stress on cellular physiology remains incomplete. We investigated the impact of ER stress on ER and inner nuclear membrane protein quality control mechanisms in Saccharomyces cerevisiae. We analyzed the turnover of substrates of four ubiquitin ligases (Doa10, Rkr1/Ltn1, Hrd1, and the Asi complex) and the metalloprotease Ste24 in induced models of ER stress. ER stress did not substantially impact Doa10 or Rkr1 substrates. However, Hrd1-mediated destruction of a protein that aberrantly engages the translocon (Deg1-Sec62) and substrates with luminal degradation signals was markedly impaired by ER stress; by contrast, Hrd1-dependent degradation of proteins with intramembrane degrons was largely unperturbed by ER stress. ER stress impaired the degradation of one of two Asi substrates analyzed and caused a translocon-clogging Ste24 substrate to accumulate in a form consistent with persistent translocon occupation. Degradation of Deg1-Sec62 in the absence of stress and stabilization during ER stress were independent of four ER stress?sensing pathways. Our results indicate ER stress differentially impacts degradation of protein quality control substrates, including those mediated by the same ubiquitin ligase. These observations suggest the existence of additional regulatory mechanisms dictating substrate selection during ER stress.

Wolbachia bacteria inhabit the cells of about half of all arthropod species, an unparalleled success stemming in large part from selfish invasive strategies. Cytoplasmic incompatibility (CI), whereby the symbiont makes itself essential to embryo viability, is the most common of these and constitutes a promising weapon against vector-borne diseases. After decades of theoretical and experimental struggle, major recent advances have been made toward a molecular understanding of this phenomenon. As pieces of the puzzle come together, from yeast and Drosophila fly transgenesis to CI diversity patterns in natural mosquito populations, it becomes clearer than ever that the CI induction and rescue stem from a toxin-antidote (TA) system. Further, the tight association of the CI genes with prophages provides clues to the possible evolutionary origin of this phenomenon and the levels of selection at play.

Numerous nascent proteins undergo folding and maturation within the luminal and membrane compartments of the endoplasmic reticulum (ER). Despite the presence of various factors in the ER that promote protein folding, many proteins fail to properly fold and assemble and are subsequently degraded. Regulatory proteins in the ER also undergo degradation in a way that is responsive to stimuli or the changing needs of the cell. As in most cellular compartments, the ubiquitin-proteasome system (UPS) is responsible for the majority of the degradation at the ER-in a process termed ER-associated degradation (ERAD). Autophagic processes utilizing ubiquitin-like protein-conjugating systems also play roles in protein degradation at the ER. The ER is continuous with the nuclear envelope (NE), which consists of the outer nuclear membrane (ONM) and inner nuclear membrane (INM). While ERAD is known also to occur at the NE, only some of the ERAD ubiquitin-ligation pathways function at the INM. Protein degradation machineries in the ER/NE target a wide variety of substrates in multiple cellular compartments, including the cytoplasm, nucleoplasm, ER lumen, ER membrane, and the NE. Here, we review the protein degradation machineries of the ER and NE and the underlying mechanisms dictating recognition and processing of substrates by these machineries.