Subdomain-specific localization of CLIMP-63 (p63) in the endoplasmic reticulum is mediated by its luminal α-helical segment

Dieter R. Klopfenstein, Judith Klumperman, Ariel Lustig, Richard A. Kammerer, Viola Oorschot, Hans Peter Hauri

    Research output: Contribution to journalArticlepeer-review


    The microtubule-binding integral 63 kD cytoskeleton-linking membrane protein (CLIMP-63; former name, p63) of the rough endoplasmic reticulum (ER) is excluded from the nuclear envelope. We studied the mechanism underlying this ER subdomain-specific localization by mutagenesis and structural analysis. Deleting the luminal but not cytosolic segment of CLIMP-63 abrogated subdomain-specific localization, as visualized by confocal microscopy in living cells and by immunoelectron microscopy using ultrathin cryosections. Photobleaching/recovery analysis revealed that the luminal segment determines restricted diffusion and immobility of the protein. The recombinant full-length luminal segment of CLIMP-63 formed α-helical 91-nm long rod-like structures as evident by circular dichroism spectroscopy and electron microscopy. In the analytical ultracentrifuge, the luminal segment sedimented at 25.7 S, indicating large complexes. The complexes most likely arose by electrostatic interactions of individual highly charged coiled coils. The findings indicate that the luminal segment of CLIMP-63 is necessary and sufficient for oligomerization into α-helical complexes that prevent nuclear envelope localization. Concentration of CLIMP-63 into patches may enhance microtubule binding on the cytosolic side and contribute to ER morphology by the formation of a protein scaffold in the lumen of the ER.
    Original languageEnglish
    Pages (from-to)1287-1299
    Number of pages12
    JournalJournal of Cell Biology
    Issue number6
    Publication statusPublished - 11 Jun 2001


    • CLIMP-63 (p63)
    • Coiled coils
    • Endoplasmic reticulum
    • Green fluorescent protein
    • Microtubule-binding membrane protein


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