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Pleckstrin Homology Domains of Phospholipase C-γ1 Directly Interact with β-Tubulin for Activation of Phospholipase C-γ1 and Reciprocal Modulation of β-Tubulin Function in Microtubule Assembly

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Author(s)
Jong-Soo ChangSung-Kuk KimTaeg-Kyu KwonSun Sik BaeDo Sik MinYoung Han LeeSoon-Ok KimJeong-Kon SeoJang Hyun ChoiPann-Ghill Suhi
Keimyung Author(s)
Kwon, Taeg Kyu
Department
Dept. of Immunology (면역학)
Journal Title
Journal of Biological Chemistry
Issued Date
2005
Volume
280
Issue
8
Abstract
Phosphoinositide-specific phospholipase C-γ1 (PLC-γ1) has two pleckstrin homology (PH) domains, an N-terminal domain and a split PH domain. Here we show that pull down of NIH3T3 cell extracts with PLC-γ1 PH domain-glutathione S-transferase fusion proteins, followed by matrix-assisted laser desorption ionization-time of flight-mass spectrometry, identified β-tubulin as a binding protein of both PLC-γ1 PH domains. Tubulin is a main component of microtubules and mitotic spindle fibers, which are composed of α- and β-tubulin heterodimers in all eukaryotic cells. PLC-γ1 and β-tubulin colocalized in the perinuclear region in COS-7 cells and cotranslocated to the plasma membrane upon agonist stimulation. Membrane-targeted translocation of depolymerized tubulin by agonist stimulation was also supported by immunoprecipitation analyses. The phosphatidylinositol 4,5-bisphosphate (PIP2) hydrolyzing activity of PLC-γ1 was substantially increased in the presence of purified tubulin in vitro, whereas the activity was not promoted by bovine serum albumin, suggesting that β-tubulin activates PLC-γ1. Furthermore, indirect immunofluorescent microscopy showed that PLC-γ1 was highly concentrated in mitotic spindle fibers, suggesting that PLC-γ1 is involved in spindle fiber formation. The effect of PLC-γ1 in microtubule formation was assessed by overexpression and silencing PLC-γ1 in COS-7 cells, which resulted in altered microtubule dynamics in vivo. Cells overexpressing PLC-γ1 showed higher microtubule densities than controls, whereas PLC-γ1 silencing with small interfering RNAs led to decreased microtubule network densities as compared with control cells. Taken together, our results suggest that PLC-γ1 and β-tubulin transmodulate each other, i.e. that PLC-γ1 modulates microtubule assembly by β-tubulin, and β-tubulin promotes PLC-γ1 activity.
Keimyung Author(s)(Kor)
권택규
Publisher
School of Medicine
Citation
Jong-Soo Chang et al. (2005). Pleckstrin Homology Domains of Phospholipase C-γ1 Directly Interact with β-Tubulin for Activation of Phospholipase C-γ1 and Reciprocal Modulation of β-Tubulin Function in Microtubule Assembly. Journal of Biological Chemistry, 280(8), 6897–6905. doi: 10.1074/jbc.M406350200
Type
Article
ISSN
0021-9258
DOI
10.1074/jbc.M406350200
URI
https://kumel.medlib.dsmc.or.kr/handle/2015.oak/33739
Appears in Collections:
1. School of Medicine (의과대학) > Dept. of Immunology (면역학)
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