Phosphorylation controls autoinhibition of cytoplasmic linker protein-170

• Published in: Molecular biology of the cell 2010-08, Vol.21 (15), p.2661-2673
• Main Authors: Lee, Ho-Sup, Komarova, Yulia A, Nadezhdina, Elena S, Anjum, Rana, Peloquin, John G, Schober, Joseph M, Danciu, Oana, van Haren, Jeffrey, Galjart, Niels, Gygi, Steven P, Akhmanova, Anna, Borisy, Gary G
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Cell Line; Cyclic AMP-Dependent Protein Kinases - metabolism; Dyneins - metabolism; Mice; Microtubule-Associated Proteins - antagonists & inhibitors; Microtubule-Associated Proteins - chemistry; Microtubule-Associated Proteins - metabolism; Microtubules - drug effects; Microtubules - metabolism; Models, Biological; Molecular Sequence Data; Mutation - genetics; Neoplasm Proteins - antagonists & inhibitors; Neoplasm Proteins - chemistry; Neoplasm Proteins - metabolism; Okadaic Acid - pharmacology; Phosphorylation - drug effects; Phosphoserine - metabolism; Protein Folding - drug effects; Protein Transport - drug effects; Proteome - metabolism
• ISSN: 1059-1524; 1939-4586
• DOI: 10.1091/mbc.e09-12-1036

Cytoplasmic linker protein (CLIP)-170 is a microtubule (MT) plus-end-tracking protein that regulates MT dynamics and links MT plus ends to different intracellular structures. We have shown previously that intramolecular association between the N and C termini results in autoinhibition of CLIP-170, thus altering its binding to MTs and the dynactin subunit p150(Glued) (J. Cell Biol. 2004: 166, 1003-1014). In this study, we demonstrate that conformational changes in CLIP-170 are regulated by phosphorylation that enhances the affinity between the N- and C-terminal domains. By using site-directed mutagenesis and phosphoproteomic analysis, we mapped the phosphorylation sites in the third serine-rich region of CLIP-170. A phosphorylation-deficient mutant of CLIP-170 displays an "open" conformation and a higher binding affinity for growing MT ends and p150(Glued) as compared with nonmutated protein, whereas a phosphomimetic mutant confined to the "folded back" conformation shows decreased MT association and does not interact with p150(Glued). We conclude that phosphorylation regulates CLIP-170 conformational changes resulting in its autoinhibition.