Binding of the Vesicle Docking Protein p115 to Golgi Membranes Is Inhibited under Mitotic Conditions*
- From the Cell Biology Laboratory, Imperial Cancer Research Fund, 44 Lincoln's Inn Fields, London WC2A 3PX, United Kingdom
- ‡ To whom correspondence should be addressed. Tel.: 44-171-269-3561; Fax: 44-171-269-3417.
Abstract
The vesicle docking protein p115 showed saturable, high affinity binding to interphase Golgi membranes. The affinity of binding was up to 20-fold lower using membranes preincubated with mitotic cytosol. In contrast, binding was not affected by mitotic pretreatment of p115. The reduction in p115 binding was mediated by phosphorylation, could be induced by a cyclin-dependent kinase, and was fully reversible. A shift of p115 from membranes to cytosol was also found after fractionating mitotic cells. The functional significance of the decreased binding was addressed by in vitro mitotic incubations which disassemble Golgi cisternae, predominantly producing transport vesicles. The addition of excess p115 decreased loss of membrane from cisternae, indicating that p115's action is limiting while transport vesicles accumulate. The cessation of intra-Golgi traffic in mitosis has been hypothesized to result from an inhibition of membrane fusion while budding of transport vesicles continues. This process also contributes to mitotic Golgi disassembly. Our results imply that there is a mitotic modification to Golgi membranes leading to a reduction in the affinity of the p115 receptor. Reduced p115 binding may play a part in the inhibition of membrane fusion by preventing prior vesicle docking.
Footnotes
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↵* The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- COP
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coat protomer
- SNARE
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SNAP receptor
- v- and t-SNAREs
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vesicle and target SNAP receptors
- SNAP
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soluble NSF attachment protein
- NSF
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N-ethylmaleimide-sensitive factor, rab-GDI, Rab-guanine nucleotide dissociation inhibitor
- PAGE
-
polyacrylamide gel electrophoresis
- MES
-
4-morpholinoethanesulfonic acid.
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- Received December 22, 1995.
- Revision received April 18, 1996.
- © 1996 by The American Society for Biochemistry and Molecular Biology, Inc.
