OST4 is a subunit of the mammalian oligosaccharyltransferase required for efficient N-glycosylation

Audrey Dumax-Vorzet, Peristera Roboti, Stephen High

    Research output: Contribution to journalArticlepeer-review

    Abstract

    The eukaryotic oligosaccharyltransferase (OST) is a membrane-embedded protein complex that catalyses the N-glycosylation of nascent polypeptides in the lumen of the endoplasmic reticulum (ER), a highly conserved biosynthetic process that enriches protein structure and function. All OSTs contain a homologue of the catalytic STT3 subunit, although in many cases this is assembled with several additional components that influence function. In S. cerevisiae, one such component is Ost4p, an extremely small membrane protein that appears to stabilise interactions between subunits of assembled OST complexes. OST4 has been identified as a putative human homologue, but to date neither its relationship to the OST complex, nor its role in protein N-glycosylation, have been directly addressed. Here, we establish that OST4 is assembled into native OST complexes containing either the catalytic STT3A or STT3B isoforms. Co-immunoprecipitation studies suggest that OST4 associates with both STT3 isoforms and with ribophorin I, an accessory subunit of mammalian OSTs. These presumptive interactions are perturbed by a single amino acid change in the transmembrane region of OST4. Using siRNA knockdowns and native gel analysis, we show that OST4 plays an important role in maintaining the stability of native OST complexes. Hence, upon OST4 depletion well-defined OST complexes are partially destabilised and a novel ribophorin I-containing subcomplex can be detected. Strikingly, cells depleted of either OST4 or STT3A show a remarkably similar defect in the N-glycosylation of endogenous prosaposin. We conclude that OST4 most likely promotes co-translational N-glycosylation by stabilising STT3A-containing OST isoforms. © 2013. Published by The Company of Biologists Ltd.
    Original languageEnglish
    Pages (from-to)2595-2606
    Number of pages11
    JournalJournal of Cell Science
    Volume126
    Issue number12
    DOIs
    Publication statusPublished - Jun 2013

    Keywords

    • Endoplasmic reticulum
    • Glycoprotein synthesis
    • Hypoglycosylation
    • Membrane protein complex
    • Ribophorin I
    • STT3 isoforms

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