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Three principal ER quality-control mechanisms, namely, the unfolded protein response, ER-associated degradation (ERAD), and ER-phagy are each important for the maintenance of ER homeostasis, yet how they are integrated to regulate ER homeostasis and organellar architecture in vivo is largely unclear. Here we report intricate crosstalk among the 3 pathways, centered around the SEL1L-HRD1 protein complex of ERAD, in the regulation of organellar organization in β cells. SEL1L-HRD1 ERAD deficiency in β cells triggers activation of autophagy, at least in part, via IRE1α (an endogenous ERAD substrate). In the absence of functional SEL1L-HRD1 ERAD, proinsulin is retained in the ER as high molecular weight conformers, which are subsequently cleared via ER-phagy. A combined loss of both SEL1L and autophagy in β cells leads to diabetes in mice shortly after weaning, with premature death by approximately 11 weeks of age, associated with marked ER retention of proinsulin and β cell loss. Using focused ion beam scanning electron microscopy powered by deep-learning automated image segmentation and 3D reconstruction, our data demonstrate a profound organellar restructuring with a massive expansion of ER volume and network in β cells lacking both SEL1L and autophagy. These data reveal at an unprecedented detail the intimate crosstalk among the 3 ER quality-control mechanisms in the dynamic regulation of organellar architecture and β cell function.

Three principal ER quality-control mechanisms, namely, the unfolded protein response, ER-associated degradation (ERAD), and ER-phagy are each important for the maintenance of ER homeostasis, yet how they are integrated to regulate ER homeostasis and organellar architecture in vivo is largely unclear. Here we report intricate crosstalk among the 3 pathways, centered around the SEL1L-HRD1 protein complex of ERAD, in the regulation of organellar organization in [beta] cells. SEL1L-HRD1 ERAD deficiency in [beta] cells triggers activation of autophagy, at least in part, via IRE1[alpha] (an endogenous ERAD substrate). In the absence of functional SEL1L-HRD1 ERAD, proinsulin is retained in the ER as high molecular weight conformers, which are subsequently cleared via ER-phagy. A combined loss of both SEL1L and autophagy in [beta] cells leads to diabetes in mice shortly after weaning, with premature death by approximately 11 weeks of age, associated with marked ER retention of proinsulin and [beta] cell loss. Using focused ion beam scanning electron microscopy powered by deep-learning automated image segmentation and 3D reconstruction, our data demonstrate a profound organellar restructuring with a massive expansion of ER volume and network in [beta] cells lacking both SEL1L and autophagy. These data reveal at an unprecedented detail the intimate cross-talk among the 3 ER quality-control mechanisms in the dynamic regulation of organellar architecture and [beta] cell function., Introduction Pancreatic [beta] cells are professional secretory cells capable of synthesizing thousands of proinsulin molecules per second in the ER (1). ER architecture and proteostasis are both highly dynamic and [...]

During entry, human papillomavirus (HPV) traffics from the endosome to the trans Golgi network (TGN) and Golgi and then the nucleus to cause infection. Although dynein is thought to play a role in HPV infection, how this host motor recruits the virus to support infection and which entry step(s) requires dynein are unclear. Here we show that the dynein cargo adaptor BICD2 binds to the HPV L2 capsid protein during entry, recruiting HPV to dynein for transport of the virus along the endosome-TGN/Golgi axis to promote infection. In the absence of BICD2 function, HPV accumulates in the endosome and TGN and infection is inhibited. Cell-based and in vitro binding studies identified a short segment near the C-terminus of L2 that can directly interact with BICD2. Our results reveal the molecular basis by which the dynein motor captures HPV to promote infection and identify this virus as a novel cargo of the BICD2 dynein adaptor. Author summary: HPV traffics from the endosome, the TGN/Golgi, and then the nucleus to cause infection, although how this is accomplished is unclear. In this manuscript, we demonstrate that the dynein cargo adaptor BICD2 interacts directly with the HPV L2 capsid protein, ferrying the virus from the endosome to the TGN/Golgi in order to promote infection. [ABSTRACT FROM AUTHOR]

Lysosomal degradation of the endoplasmic reticulum (ER) and its components through the autophagy pathway has emerged as a major regulator of ER proteostasis. Commonly referred to as ER‐phagy and ER‐to‐lysosome‐associated degradation (ERLAD), how the ER is targeted to the lysosome has been recently clarified by a growing number of studies. Here, we summarize the discoveries of the molecular components required for lysosomal degradation of the ER and their proposed mechanisms of action. Additionally, we discuss how cells employ these machineries to create the different routes of ER‐lysosome‐associated degradation. Further, we review the role of ER‐phagy in viral infection pathways, as well as the implication of ER‐phagy in human disease. In sum, we provide a comprehensive overview of the current field of ER‐phagy. [ABSTRACT FROM AUTHOR]

Both basal and glucose-stimulated insulin release occur primarily by insulin secretory granule exocytosis from pancreatic [beta] cells, and both are needed to maintain normoglycemia. Loss of insulin-secreting [beta] cells, accompanied by abnormal glucose tolerance, may involve simple exhaustion of insulin reserves (which, by immunostaining, appears as a loss of [beta] cell identity), or [beta] cell dedifferentiation, or [beta] cell death. While various sensing and signaling defects can result in diminished insulin secretion, somewhat less attention has been paid to diabetes risk caused by insufficiency in the biosynthetic generation and maintenance of the total insulin granule storage pool. This Review offers an overview of insulin biosynthesis, beginning with the preproinsulin mRNA (translation and translocation into the ER), proinsulin folding and export from the ER, and delivery via the Golgi complex to secretory granules for conversion to insulin and ultimate hormone storage. All of these steps are needed for generation and maintenance of the total insulin granule pool, and defects in any of these steps may, weakly or strongly, perturb glycemic control. The foregoing considerations have obvious potential relevance to the pathogenesis of type 2 diabetes and some forms of monogenic diabetes; conceivably, several of these concepts might also have implications for [beta] cell failure in type 1 diabetes., Brief description of insulin mRNA quantity and quality Many factors may contribute to the control of insulin mRNA levels (1, 2), including transcriptional networks that control [beta] cell function (3) [...]

The multi-functional endoplasmic reticulum (ER) is exploited by viruses to cause infection. Morphologically, this organelle is a highly interconnected membranous network consisting of sheets and tubules whose levels are dynamic, changing in response to cellular conditions. Functionally, the ER is responsible for protein synthesis, folding, secretion and degradation, as well as Ca2+ homeostasis and lipid biosynthesis, with each event catalyzed by defined ER factors. Strikingly, these ER host factors are hijacked by viruses to support different infection steps, including entry, translation, replication, assembly and egress. Although the full repertoire of these ER factors that are hijacked is unknown, recent studies have uncovered several ER membrane machineries that are exploited by viruses - ranging from polyomavirus to flavivirus and coronavirus - to facilitate different steps of their life cycle. These discoveries should provide better understanding of virus infection mechanisms, potentially leading to the development of more effective anti-viral therapies. [ABSTRACT FROM AUTHOR]

Misfolded proteins in the endoplasmic reticulum (ER) are retained in the organelle or retrotranslocated to the cytosol for proteasomal degradation. ER chaperones that guide these opposing processes are largely unknown. We developed a semipermeabilized cell system to study the retrotranslocation of cholera toxin (CT), a toxic agent that crosses the ER membrane to reach the cytosol during intoxication. We found that protein disulfide isomerase (PDI) facilitates CT retrotranslocation, whereas ERp72, a PDI-like protein, mediates its ER retention. In vitro analysis revealed that PDI and ERp72 alter CT's conformation in a manner consistent with their roles in retrotranslocation and ER retention. Moreover, we found that PDI's and ERp72's opposing functions operate on endogenous ER misfolded proteins. Thus, our data identify PDI family proteins that play opposing roles in ER quality control and establish an assay to further delineate the mechanism of CT retrotranslocation.

Cholera toxin (CT) and related A[B.sub.5]-subunit toxins move from the plasma membrane through the trans-Golgi and endoplasmic reticulum (ER) to the cytosol of host cells. The toxins exploit a specific glycolipid pathway rather than a protein pathway. They bind glycolipids that associate with lipid rafts at the cell surface, which carry the toxins retrograde to the Golgi and ER. In the ER, the A1-chain of the CT unfolds and enters the cytosol by hijacking the cellular machinery that enables misfolded proteins to cross the membrane for degradation by the proteasome, a process termed retro-translocation. Upon entering the cytosol, the Al-chain rapidly refolds, avoids the proteasome and induces toxicity.

Protein disulfide isomerase (PDI) has been found to act as a redox-dependent chaperone in the unfolding of cholera toxin in the endoplasmic reticulum (ER) lumen. This happens after the A chain of the toxin has been cleaved. A role for PDI is suggested in retrograde protein transport into the cytosol, and it can function as a novel type of chaperone. The binding and release of substrates would then be regulated by a redox cycle, not an ATPase cycle.

ER-specific autophagy (reticulophagy) has emerged as a critical degradative route for misfolded secretory proteins. Our previous work showed that RTN3 (reticulon 3) drives reticulophagic clearance of disease-causing mutant prohormones. How RTN3, a protein residing on the cytosolic leaflet of the ER bilayer, recruits these lumenally-localized cargos has remained a mystery. To address this question, we used an unbiased proteomics approach to identify RTN3-interacting partners. We discovered that RTN3 recruits misfolded prohormones for lysosomal degradation through the ER transmembrane protein PGRMC1. RTN3 complexes with PGRMC1, which directly binds to misfolded prohormones via its distal ER lumenal domain. Cargos for the RTN3-PGRMC1 degradative axis include mutant POMC (proopiomelanocortin) and proinsulin, each of which oligomerizes in the ER during misfolding, entrapping their wild-type counterparts, leading to secretion defects. Although reticulophagy is thought to degrade large protein aggregates, PGRMC1 instead selectively recruits and promotes degradation of only small oligomers of the mutant prohormones. Of physiological importance, genetic or pharmacological inactivation of PGRMC1 in pancreatic β-cells expressing both wild-type and mutant proinsulin impairs mutant proinsulin turnover and promotes trafficking of wild-type proinsulin. These findings pinpoint PGRMC1 as a possible intervention point for diseases caused by ER protein retention. [ABSTRACT FROM AUTHOR]

The endoplasmic reticulum (ER), with its expansive membranous system and a vast network of chaperones, enzymes, sensors, and ion channels, orchestrates diverse cellular functions, ranging from protein synthesis, folding, secretion, and degradation to lipid biogenesis and calcium homeostasis. Strikingly, some of the functions of the ER are exploited by viruses to promote their life cycles. During entry, viruses must penetrate a host membrane and reach an intracellular destination to express and replicate their genomes. These events lead to the assembly of new viral progenies that exit the host cell, thereby initiating further rounds of infection. In this review, we highlight how three distinct viruses -- polyomavirus, flavivirus, and coronavirus -- co-opt key functions of the ER to cause infection. We anticipate that illuminating this virus-ER interplay will provide rational therapeutic approaches to combat the virus-induced diseases. [ABSTRACT FROM AUTHOR]