| Co-Investigator(Kenkyū-buntansha) |
KOHAMA Kazuhiro Gunma University, Graduate School of Medicine, Professor, 大学院・医学系研究科, 教授 (30101116)
KUMAGAI Hiroyuki Gunma University, Graduate School of Medicine, Research Associate, 大学院・医学系研究科, 助手 (20321945)
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| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2004: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 2003: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Research Abstract |
We examined the distribution and dynamics of actin-binding proteins in Ng108-15 cells. EGFP/fascin localized in filopodia. Pseudo-unphosphorylated mutant, in which 39-serine of fascin was replaced to asparatic acid, localized in filopodia while pseudo-phosphorylated mutant, in which 39-serine was replaced to alanine, showed dispersed distribution, suggesting that localization of fascin is regulated by phosphorylation. EGFP/smooth muscle type tropomyosin localized only in stress fibers, while EGFP/nonmusle type tropomyosin localized in stress fibers, filopodia, and lamellipodia. EGFP/caldesmon localized in all the actin structures.
Reconstituted characterization of purified proteins revealed that fascin formed tight F-actin bundles with polarity. Drebrin bound to F-actin with a stoichiometry of 5 actin versus 1 drebrin molecule. Drebrin inhibited the actin-binding of caldesmon, actin-binding of tropomyosin, and actin-binding of fascin.
These results suggest that competitive binding between drebrin and tropomyosin/caldesmon, between drebrin and fascin, and between fascin and tropomyosin/caldesmon may cause the differential localization fascin, drebrin, and tropomyosin/caldesmon, resulting in different organization of actin structure.
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