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Originally published as MBC in Press, 10.1091/mbc.E03-11-0822 on July 14, 2004

Vol. 15, Issue 9, 4289-4298, September 2004

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Cdc2 Kinase-dependent Disassembly of Endoplasmic Reticulum (ER) Exit Sites Inhibits ER-to-Golgi Vesicular Transport during Mitosis{boxd}{boxv}

Fumi Kano *, Arowu R. Tanaka *, Shinobu Yamauchi *, Hisao Kondo {dagger} {ddagger}, and Masayuki Murata * §

* Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Tokyo 153-8902, Japan; {dagger} Cambridge Institute for Medical Research, University of Cambridge, Cambridge CB2 2XY, United Kingdom; and {ddagger} PRESTO, Japan Science and Technology Corporation, Saitama 332-0012, Japan

Submitted November 17, 2003; Accepted June 30, 2004
Monitoring Editor: Benjamin Glick

We observed the disassembly of endoplasmic reticulum (ER) exit sites (ERES) by confocal microscopy during mitosis in Chinese hamster ovary (CHO) cells by using Yip1A fused to green fluorescence protein (GFP) as a transmembrane marker of ERES. Photobleaching experiments revealed that Yip1A-GFP, which was restricted to the ERES during interphase, diffused throughout the ER network during mitosis. Next, we reconstituted mitotic disassembly of Yip1A-GFP–labeled ERES in streptolysin O-permeabilized CHO cells by using mitotic L5178Y cytosol. Using the ERES disassembly assay and the anterograde transport assay of GFP-tagged VSVGts045, we demonstrated that the phosphorylation of p47 by Cdc2 kinase regulates the disassembly of ERES and results in the specific inhibition of ER-to-Golgi transport during mitosis.

Article published online ahead of print. Mol. Biol. Cell 10.1091/mbc.E03–11–0822. Article and publication date are available at www.molbiolcell.org/cgi/doi/10.1091/mbc.E03–11–0822.

Abbreviations used: ERES, endoplasmic reticulum exit site(s); NEM, N-ethylmaleimide; SLO, streptolysin O, TB, transport buffer; TC, transport complex(es); VSVG, vesicular stomatitis virus G protein.

{boxd}{boxv} Online version of this article contains supporting material. Online version is available at www.molbiolcell.org.

§ Corresponding author. E-mail address: mmurata{at}bio.c.u-tokyo.ac.jp.




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