Elimination of excess smooth endoplasmic reticulum after phenobarbital administration.

Feldman, Dorothy; Swarm, Richard L.; Becker, J. M.

doi:10.1177/28.9.7410819

Cited by 41 publications

(23 citation statements)

References 21 publications

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“…Subsequently, cells were permeabilized with 0.25% saponin, 0.1% BSA and blocked in blocking buffer (0.2% BSA, 5% goat serum, 50 mM NH 4 Cl, 0.1% saponin, 20 mM PO 4 buffer, 150 mM NaCl). Staining with primary antibodies and nanogold labeled secondary antibodies was performed in blocking buffer.…”

Section: Immunogold Electron Microscopymentioning

confidence: 99%

Translocon component Sec62 acts in endoplasmic reticulum turnover during stress recovery

et al. 2016

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The endoplasmic reticulum (ER) is a site of protein biogenesis in eukaryotic cells. Perturbing ER homeostasis activates stress programs collectively called the unfolded protein response (UPR). The UPR enhances production of ER-resident chaperones and enzymes to reduce the burden of misfolded proteins. On resolution of ER stress, ill-defined, selective autophagic programs remove excess ER components. Here we identify Sec62, a constituent of the translocon complex regulating protein import in the mammalian ER, as an ER-resident autophagy receptor. Sec62 intervenes during recovery from ER stress to selectively deliver ER components to the autolysosomal system for clearance in a series of events that we name recovER-phagy. Sec62 contains a conserved LC3-interacting region in the C-terminal cytosolic domain that is required for its function in recovER-phagy, but is dispensable for its function in the protein translocation machinery. Our results identify Sec62 as a critical molecular component in maintenance and recovery of ER homeostasis. DOI: https://doi.org/10.1038/ncb3423Posted at the Zurich Open Repository and Archive, University of Zurich ZORA URL: https://doi.org/10.5167/uzh-127515 Accepted Version Originally published at: Fumagalli, Fiorenza; Noak, Julia; Bergmann, Timothy J; Presmanes, Eduardo Cebollero; Pisoni, Giorgia Brambilla; Fasana, Elisa; Fregno, Ilaria; Galli, Carmela; Loi, Marisa; Solda, Tatiana; D'Antuono, Rocco; Raimondi, Andrea; Jung, Martin; Melnyk, Armin; Schorr, Stefan; Schreiber, Anne; Simonelli, Luca; Varani, Luca; Wilson-Zbinden, Caroline; Zerbe, Oliver; Hofmann, Kay; Peter, Matthias; Quadroni, Manfredo; Zimmermann, Richard; Molinari, Maurizio (2016 To define mechanisms that regulate the return of ER-resident chaperones and folding factors to their physiologic intracellular level after resolution of an ER stress, we established a protocol for reversible induction of UPR in cultured mammalian cells (Fig. 1a). Briefly, human embryonic kidney cells (HEK293) or mouse embryonic fibroblasts (MEF) were exposed for 12 h to non-toxic doses of cyclopiazonic acid (CPA), a reversible inhibitor of the sarco/endoplasmic reticulum calcium pump 6 . The return of ER-resident gene products at their pre-stress level was monitored during resolution of the UPR obtained upon CPA wash out ( CPA wash out initiated a recovery phase characterized by the rapid return of ER stress-induced transcripts at, or below, their pre-stress levels (Fig. 1b, recovery, T 1/2 average ≈ 1 h, blue line). The corresponding ER stress-induced proteins returned to their physiologic levels with much slower kinetics (Fig. 1c, d, T 1/2 average ≈ 10 h, blue). 3With the exception of Herp, which is rapidly turned over with intervention of proteasomes (Fig. 1c, d (Fig. 1g, 2a) and other membrane and luminal ER marker proteins such as Sec62 and Crt ( Fig. 2b and Extended data Fig. 3) in 0.5-1.5 µm diameter cytoplasmic puncta that rapidly disappeared upon BafA1 wash out (Extended data Fig. 4). Cytosolic puncta containing ER marker prot...

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Section: Immunogold Electron Microscopymentioning

confidence: 99%

Translocon component Sec62 acts in endoplasmic reticulum turnover during stress recovery

et al. 2016

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“…In the present electron microscopic serial section analysis, no evidence for autophagic removal of EDEM1-containing parts of the ER was found. Thus, autophagy of EDEM1 comprises the clearance of cytosolic EDEM1 aggregates and has to be distinguished from the autophagy of ER cisternae containing aggregated misfolded protein [45,46], or from ER-phagy, which is the selective autophagy of surplus ER under non-steady state conditions [31,47,48]. There is other evidence that the autophagylysosome system, but not proteasomes, are involved in Figure 7.…”

Section: Discussionmentioning

confidence: 99%

Basal autophagy is involved in the degradation of the ERAD component EDEM1

Fourn

Gaplovská-Kyselá

Guhl

et al. 2009

Cell. Mol. Life Sci.

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Little is known about the fate of machinery proteins of the protein quality control and endoplasmic reticulum(ER)-associated degradation (ERAD). We investigated the degradation of the ERAD component EDEM1, which directs overexpressed misfolded glycoproteins to degradation. Endogenous EDEM1 was studied since EDEM1 overexpression not only resulted in inappropriate occurrence throughout the ER but also caused cytotoxic effects. Proteasome inhibitors had no effect on the clearance of endogenous EDEM1 in non-starved cells. However, EDEM1 could be detected by immunocytochemistry in autophagosomes and biochemically in LC3 immuno-purified autophagosomes. Furthermore, influencing the lysosome-autophagy pathway by vinblastine or pepstatin A/E64d and inhibiting autophagosome formation by 3-methyladenine or ATGs short interfering RNA knockdown stabilized EDEM1. Autophagic degradation involved removal of cytosolic Triton X-100-insoluble deglycosylated EDEM1, but not of EDEM1-containing ER cisternae. Our studies demonstrate that endogenous EDEM1 in cells not stressed by the expression of a transgenic misfolded protein reaches the cytosol and is degraded by basal autophagy.

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“…Phenobarbital induction ofcytochrome P450 over prolonged time periods in experimental animals produces major morphological and biochemical alterations in liver, including tumor formation (24)(25)(26). The marked depletion of hepatic cytochrome P-450 produced by Co-heme during the 3-month experiment (Fig.…”

Section: Discussionmentioning

confidence: 99%

The cytochrome P-450-depleted animal: an experimental model for in vivo studies in chemical biology.

Drummond¹,

Kappas²

1982

Proc. Natl. Acad. Sci. U.S.A.

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An experimental method is described to deplete markedly in vivo the cytochrome P450 content of liver for prolonged periods of time. The method uses the synthetic metalloporphyrin cobalt-heme (cobalt protoporphyrin IX), which possesses the dual biological properties ofrepressing 6-aminolevulinate synthase, the rate-limiting enzyme of heme biosynthesis, and of potently inducing microsomal heme oxygenase, the rate-limiting enzyme of heme catabolism. A single dose of cobalt-heme (125 ,umol/kg of body weight) decreased within 48 hr hepatic cytochrome P-450 to z20% of normal, at which level it remained for 10 days; normal levels were not achieved by 36 days. Periodic administration (total, six injections) of a smaller dose of cobaltheme (50 ,umol/kg of body weight) maintained the cytochrome P-450 content at levels =15% of normal for >90 days with concurrent profound impairment of mono-oxygenase reactions catalyzed by this heme protein. The ability of cobalt-heme to produce profound and prolonged depletion of cytochrome P-450 in vivo provides a valuable model for examining the role ofcytochrome P-450-dependent metabolism in the biology of endogenous and exogenous chemicals. MATERIALS AND METHODS Materials. Male Sprague-Dawley rats (175-200 g) purchased from Holtzman, (Madison, WI) were used throughout this study. Rats were injected subcutaneously with single doses of Co-heme, up to 125 ,umoVkg of body weight unless otherwise indicated. The animals were starved for 16 hr but permitted access to water prior to sacrifice. Solutions of Co-heme were prepared by dissolving the compound in a small volume (0.1 mV ml of Co-heme solution) of 0.1 M sodium hydroxide, adjusting the pH to 7.4 with 1 M HC1, and making up to final volume with 0.9% NaCl. These Co-heme solutions were administered (0.2-0.4 mV/100 g ofbody weight) within 10 min ofpreparation.Control animals received an equivalent volume of 0.9% NaCl. In the chronic-condition experiment, animals received six doses of Co-heme (50 ,umoVkg of body weight, each dose), the first on day 0 of the study and the subsequent ones on days 7, 14, 32, 60, and 88.Co-heme was supplied by Porphyrin Products (Logan, UT). All other chemicals used were of the highest grade obtainable and were supplied by either Sigma or Fisher.Tissue Preparation. Livers were perfused in situ with icecold 0.9% NaCl, removed, and homogenized, as were other tissues, in 3 vol of 0.1 M potassium phosphate, pH 7.4/0.25 M sucrose. The homogenate was centrifuged at 9000 X g for 20 min. The precipitate was washed twice with 0.1 M potassium phosphate, pH 7.4, prior to assaying for AmLev synthase activity. The 9000 X g supernatant was centrifuged at 100,000x g for 60 min; the cytosol from untreated liver was used as a source of biliverdin reductase and the microsomal pellet was washed once with and then resuspended in 0.1 M potassium phosphate, pH 7.4, at a protein concentration of 15-20 mg/ml.Enzyme Assays. The activities of AmLev synthase, aniline hydroxylase, ethylmorphine demethylase, aryl hydrocarbon hydro...

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Elimination of excess smooth endoplasmic reticulum after phenobarbital administration.

Cited by 41 publications

References 21 publications

Translocon component Sec62 acts in endoplasmic reticulum turnover during stress recovery

Translocon component Sec62 acts in endoplasmic reticulum turnover during stress recovery

Basal autophagy is involved in the degradation of the ERAD component EDEM1

The cytochrome P-450-depleted animal: an experimental model for in vivo studies in chemical biology.

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