Journal
JOURNAL OF MOLECULAR BIOLOGY
Volume 395, Issue 5, Pages 1049-1062Publisher
ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD
DOI: 10.1016/j.jmb.2009.11.014
Keywords
microtubule; CLIP-170; EB1; +TIP; tubulin
Categories
Funding
- National Institutes of Health [R01 GM065420]
- American Heart Association [0825871G]
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Cytoplasmic linker protein 170 (CLIP-170) is a microtubule (MT) plus-end tracking protein (+TIP) that dynamically localizes to the MT plus end and regulates MT dynamics. The mechanisms of these activities remain unclear because the CLIP-170-MT interaction is poorly understood, and even less is known about how CLIP-170 and other +TIPs act together as a network. CLIP-170 binds to the acidic C-terminal tail of alpha-tubulin. However, the observation that CLIP-170 has two CLIP-Gly (cytoskeleton-associated protein glycine-rich) motifs and multiple serine-rich regions Suggests that a single CLIP-170 molecule has multiple tubulin binding sites, and that these sites might bind to multiple parts of the tubulin dimer. Using a combination of chemical cross-linking and mass spectrometry, we find that CLIP-170 binds to both alpha-tubulin and beta-tubulin, and that binding is not limited to the acidic C-terminal tails. We provide evidence that these additional binding sites include the H12 helices of both alpha-tubulin and tubulin and are significant for CLIP-170 activity. Previous work has shown that CLIP-170 binds to end-binding protein 1 (EB1) via the EB1 C-terminus, which mimics the acidic C-terminal tail of tubulin. We find that CLIP-170 can utilize its multiple tubulin binding sites to bind to EB1 and MT simultaneously. These observations help to explain how CLIP-170 can nucleate MTs and alter MT dynamics, and they contribute to understanding the significance and properties of the +TIP network. (C) 2009 Elsevier Ltd All rights reserved.
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