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J Cell Biol. 2008 Feb 25;180(4):729-37. doi: 10.1083/jcb.200707203. Epub 2008 Feb 18.

Dynamic behavior of GFP-CLIP-170 reveals fast protein turnover on microtubule plus ends.

Author information

1
Department of Cell Biology and Genetics, Erasmus Medical Center, 3000 DR Rotterdam, Netherlands.

Abstract

Microtubule (MT) plus end-tracking proteins (+TIPs) specifically recognize the ends of growing MTs. +TIPs are involved in diverse cellular processes such as cell division, cell migration, and cell polarity. Although +TIP tracking is important for these processes, the mechanisms underlying plus end specificity of mammalian +TIPs are not completely understood. Cytoplasmic linker protein 170 (CLIP-170), the prototype +TIP, was proposed to bind to MT ends with high affinity, possibly by copolymerization with tubulin, and to dissociate seconds later. However, using fluorescence-based approaches, we show that two +TIPs, CLIP-170 and end-binding protein 3 (EB3), turn over rapidly on MT ends. Diffusion of CLIP-170 and EB3 appears to be rate limiting for their binding to MT plus ends. We also report that the ends of growing MTs contain a surplus of sites to which CLIP-170 binds with relatively low affinity. We propose that the observed loss of fluorescent +TIPs at plus ends does not reflect the behavior of single molecules but is a result of overall structural changes of the MT end.

PMID:
18283108
PMCID:
PMC2265578
DOI:
10.1083/jcb.200707203
[Indexed for MEDLINE]
Free PMC Article

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