Co-chaperone Specificity in Gating of the Polypeptide Conducting Channel in the Membrane of the Human Endoplasmic Reticulum |
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Authors: | Stefan Schorr Marie-Christine Klein Igor Gamayun Armin Melnyk Martin Jung Nico Sch?uble Qian Wang Birgit Hemmis Florian Bochen Markus Greiner Pavel Lampel Sabine Katharina Urban Sarah Hassdenteufel Johanna Dudek Xing-Zhen Chen Richard Wagner Adolfo Cavalié Richard Zimmermann |
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Institution: | From the Departments of ‡Medical Biochemistry and Molecular Biology and ;§Experimental and Clinical Pharmacology and Toxicology, Saarland University, 66421 Homburg, Germany, ;the ¶Department of Physiology, University of Alberta, Edmonton T6G 2H7, Canada, and ;the ‖Division of Biophysics, Universität Osnabrück, FB Biologie/Chemie, 49076 Osnabrück, Germany |
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Abstract: | In mammalian cells, signal peptide-dependent protein transport into the endoplasmic reticulum (ER) is mediated by a dynamic polypeptide-conducting channel, the heterotrimeric Sec61 complex. Previous work has characterized the Sec61 complex as a potential ER Ca2+ leak channel in HeLa cells and identified ER lumenal molecular chaperone immunoglobulin heavy-chain-binding protein (BiP) as limiting Ca2+ leakage via the open Sec61 channel by facilitating channel closing. This BiP activity involves binding of BiP to the ER lumenal loop 7 of Sec61α in the vicinity of tyrosine 344. Of note, the Y344H mutation destroys the BiP binding site and causes pancreatic β-cell apoptosis and diabetes in mice. Here, we systematically depleted HeLa cells of the BiP co-chaperones by siRNA-mediated gene silencing and used live cell Ca2+ imaging to monitor the effects on ER Ca2+ leakage. Depletion of either one of the ER lumenal BiP co-chaperones, ERj3 and ERj6, but not the ER membrane-resident co-chaperones (such as Sec63 protein, which assists BiP in Sec61 channel opening) led to increased Ca2+ leakage via Sec6 complex, thereby phenocopying the effect of BiP depletion. Thus, BiP facilitates Sec61 channel closure (i.e. limits ER Ca2+ leakage) via the Sec61 channel with the help of ERj3 and ERj6. Interestingly, deletion of ERj6 causes pancreatic β-cell failure and diabetes in mice and humans. We suggest that co-chaperone-controlled gating of the Sec61 channel by BiP is particularly important for cells, which are highly active in protein secretion, and that breakdown of this regulatory mechanism can cause apoptosis and disease. |
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Keywords: | calcium imaging chaperone diabetes endoplasmic reticulum (ER) protein translocation BiP ER calcium leakage Sec61 complex gating cellular calcium homeostasis |
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