Cells Deploy a Two-Pronged Strategy to Rectify Misfolded Proinsulin Aggregates

Cunningham, Corey N.; Williams, Jeffrey M.; Knupp, Jeffrey; Arunagiri, Anoop; Arvan, Peter; Tsai, Billy

doi:10.1016/j.molcel.2019.05.011

Cited by 68 publications

(97 citation statements)

References 50 publications

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“…5 b); the large puncta represent Akita aggregates that can also be generated by RTN3 knockdown (Cunningham et al, 2019). These findings support the idea that, in contrast to NHK, Akita forms large aggregated complexes that are disposed of by the autophagy pathway (Cunningham et al, 2019). This notion is further supported by sucrose gradient analysis, demonstrating that in cells depleted of RTN3, Akita exists as a massive protein complex similar to purified SV40, whereas NHK fractionates as a significantly smaller protein (Fig.…”

Section: Rtn3 Protects Er Membrane Integrity During Er Exit Of Misfolsupporting

confidence: 76%

Reticulon protects the integrity of the ER membrane during ER escape of large macromolecular protein complexes

Chen

Williams

Arvan

et al. 2020

Journal of Cell Biology

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Escape of large macromolecular complexes from the endoplasmic reticulum (ER), such as a viral particle or cellular aggregate, likely induces mechanical stress initiated on the luminal side of the ER membrane, which may threaten its integrity. How the ER responds to this threat remains unknown. Here we demonstrate that the cytosolic leaflet ER morphogenic protein reticulon (RTN) protects ER membrane integrity when polyomavirus SV40 escapes the ER to reach the cytosol en route to infection. SV40 coopts an intrinsic RTN function, as we also found that RTN prevents membrane damage during ER escape of a misfolded proinsulin aggregate destined for lysosomal degradation via ER-phagy. Our studies reveal that although ER membrane integrity may be threatened during ER escape of large macromolecular protein complexes, the action of RTN counters this, presumably by deploying its curvature-inducing activity to provide membrane flexibility and stability to limit mechanical stress imposed on the ER membrane.

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Section: Rtn3 Protects Er Membrane Integrity During Er Exit Of Misfolsupporting

confidence: 76%

Reticulon protects the integrity of the ER membrane during ER escape of large macromolecular protein complexes

Chen

Williams

Arvan

et al. 2020

Journal of Cell Biology

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“…We have recently reported that the combination of upregulated AMPK activation with downregulated TOR activation is a cell survival feature in response to ER stress, conserved down to yeast (37). AMPK is an important activator of ERphagy and mitochondrial function, which is known to be stimulated in ER-stressed cells and can promote disposal of ER protein aggregates (38,39). Thus perhaps these responses are to be expected, yet to us it is particularly notable that along with decreased levels of phospho-S6-kinase, thyrocytes adapted to chronic continuous ER stress maintain a lower overall protein synthesis level while exhibiting a growth rate that ultimately is comparable to that of unstressed cells.…”

Section: Discussionmentioning

confidence: 99%

Thyrocyte cell survival and adaptation to chronic endoplasmic reticulum stress due to misfolded thyroglobulin

Morishita

Kabil

Young

et al. 2020

Journal of Biological Chemistry

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The large secretory glycoprotein thyroglobulin is the primary translation product of thyroid follicular cells. This difficult-to-fold protein is susceptible to structural alterations that disable export of the misfolded thyroglobulin from the endoplasmic reticulum (ER), which is a known cause of congenital hypothyroidism characterized by severe chronic thyrocyte ER stress. Nevertheless, individuals with this disease commonly grow a goiter, indicating thyroid cell survival and adaptation. To model these processes, here we continuously exposed rat PCCL3 thyrocytes to tunicamycin, which causes a significant degree of ER stress that is specifically attributable to thyroglobulin misfolding. We found that, in response, PCCL3 cells down-regulate expression of the “tunicamycin transporter” (major facilitator superfamily domain containing-2A, Mfsd2a). Following CRISPR/Cas9-mediated Mfsd2a deletion, PCCL3 cells could no longer escape the chronic effects of high-dose tunicamycin, as demonstrated by persistent accumulation of unglycosylated thyroglobulin; nevertheless, these thyrocytes survived and grew. A proteomic analysis of these cells adapted to chronic ER protein misfolding revealed many hundreds of up-regulated proteins, indicating stimulation of ER chaperones, oxidoreductases, stress responses, and lipid biosynthesis pathways. Further, we noted increased phospho–AMP-kinase, suggesting up-regulated AMP-kinase activity, and decreased phospho–S6-kinase and protein translation, suggesting decreased mTOR activity. These changes are consistent with conserved cell survival/adaptation pathways. We also observed a less-differentiated thyrocyte phenotype with decreased PAX8, FOXE1, and TPO protein levels, along with decreased thyroglobulin mRNA levels. In summary, we have developed a model of thyrocyte survival and growth during chronic continuous ER stress that recapitulates features of congenital hypothyroid goiter caused by mutant thyroglobulin.

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“…Indeed, RTN3 has the capacity to induce ER fragmentation via the presence of a RHD and to deliver ER portions to the autophagosomes via six LIR domains . In addition to induce tubular ER‐phagy during starvation, RTN3 is involved in the selective cargo removal of a mutant type of proinsulin ( Akita mutant) . Curiously, the mechanism of clearance of proinsulin via ER‐phagy seems to be partially independent of the LIR domain .…”

Section: The ‘Er‐phagy’ Pathwaymentioning

confidence: 99%

“…In addition to induce tubular ER‐phagy during starvation, RTN3 is involved in the selective cargo removal of a mutant type of proinsulin ( Akita mutant) . Curiously, the mechanism of clearance of proinsulin via ER‐phagy seems to be partially independent of the LIR domain . Proinsulin is composed of two chains, A and B, and the Akita mutant is characterized by a cysteine‐to‐tyrosine substitution on the A chains, leaving the B chain unpaired thus forming non‐native sulphide bonds .…”

Section: The ‘Er‐phagy’ Pathwaymentioning

confidence: 99%

Emerging lysosomal pathways for quality control at the endoplasmic reticulum

2019

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Protein misfolding occurring in the endoplasmic reticulum (ER) might eventually lead to aggregation and cellular distress, and is a primary pathogenic mechanism in multiple human disorders. Mammals have developed evolutionary‐conserved quality control mechanisms at the level of the ER. The best characterized is the ER‐associated degradation (ERAD) pathway, through which misfolded proteins translocate from the ER to the cytosol and are subsequently proteasomally degraded. However, increasing evidence indicates that additional quality control mechanisms apply for misfolded ER clients that are not eligible for ERAD. This review focuses on the alternative, ERAD‐independent, mechanisms of clearance of misfolded polypeptides from the ER. These processes, collectively referred to as ER‐to‐lysosome–associated degradation, involve ER‐phagy, microautophagy or vesicular transport. The identification of the underlying molecular mechanisms is particularly important for developing new therapeutic approaches for human diseases associated with protein aggregate formation.

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Cells Deploy a Two-Pronged Strategy to Rectify Misfolded Proinsulin Aggregates

Cited by 68 publications

References 50 publications

Reticulon protects the integrity of the ER membrane during ER escape of large macromolecular protein complexes

Reticulon protects the integrity of the ER membrane during ER escape of large macromolecular protein complexes

Thyrocyte cell survival and adaptation to chronic endoplasmic reticulum stress due to misfolded thyroglobulin

Emerging lysosomal pathways for quality control at the endoplasmic reticulum

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