Peptide-based interactions with calnexin target misassembled membrane proteins into endoplasmic reticulum-derived multilamellar bodies

JOURNAL OF MOLECULAR BIOLOGY

卷:378

Article

Korkhov, Vladimir M.^1,2  Milan-Lobo, Laura¹  Zuber, Benoit²  Farhan, Hesso¹  Schmid, Johannes A.³  Freissmuth, Michael¹  Sitte, Harald H.¹ 

[1] Med Uni Vienna, Inst Pharmacol, Ctr Biomol Med & Pharmacol, A-1090 Vienna, Austria

[2] Med Uni Vienna, Inst Vasc Biol, Ctr Biomol Med & Pharmacol, A-1090 Vienna, Austria

[3] MRC, Mol Biol Lab, Cambridge CB2 2QH, England

关键词: calnexin;    neurotransmitter transporters;    GABA transporter-1;    G protein-coupled receptors;    transmembrane domains;   

DOI  :  10.1016/j.jmb.2008.02.056

来源: Elsevier

【 摘 要 】

Oligomeric assembly of neurotransmitter transporters is a prerequisite for their export from the endoplasmic reticulum (ER) and their subsequent delivery to the neuronal synapse. We previously identified mutations, e.g., in the gamma-aminobutyric acid (GABA) transporter-1 (GAT1), which disrupted assembly and caused retention of the transporter in the ER. Using one representative mutant, GAT1-E101D, we showed here that ER retention was due to association of the transporter with the ER chaperone calnexin: interaction with calnexin led to accumulation of GAT1 in concentric bodies corresponding to previously described multilamellar ER-derived structures. The transmembrane domain of calnexin was necessary and sufficient to direct the protein into these concentric bodies. Both yellow fluorescent protein-tagged versions of wild-type GAT1 and of the GAT1-E101D mutant remained in disperse (i.e., non-aggregated) form in these concentric bodies, because fluorescence recovered rapidly (t(1/2) similar to 500 ms) upon photobleaching. Fluorescence energy resonance transfer microscopy was employed to visualize a tight interaction of GAT1-E101D with calnexin. Recognition by calnexin occurred largely in a glycan-independent manner and, at least in part, at the level of the transmembrane domain. Our findings are consistent with a model in which the transmembrane segment of calnexin participates in chaperoning the inter- and intramolecular arrangement of hydrophobic segment in oligomeric proteins. (c) 2008 Elsevier Ltd. All rights reserved.

【 授权许可】

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