| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2006: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2005: ¥2,400,000 (Direct Cost: ¥2,400,000)
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| Research Abstract |
Src-family tyrosine kinases are cytosolic enzymes, and lipid modification at their N-termini leads to their localization to endomembranes as well as the plasma membrane. In this study, we examined intracellular trafficking, localization and function of Src-family tyrosine kinases and their negative regulator Chk tyrosine kinase. Our findings are described as follows.
1. Stimulation of cells with H_2O_2 induces tyrosine phosphorylation of annexin 11 by Lyn that is present on the Golgi apparatus, leading to the translocation of annexin 11 from the Golgi to the endoplasmic reticulum.
2. Lyn kinase domain, which is important for Lyn trafficking, associates with two proteins that are identified by mass spectrometry.
3. Nuclear localization of Chk induces dynamic multi-lobulation of the nucleus and prolonged S phase of the cell cycle. The N-terminal domain of Chk associates with the nucleus and the nuclear matrix, and nuclear tyrosine phosphorylation induced by Chk contributes to inhibition of cell proliferation.
4. c-Src, unlike Lyn, c-Yes and Fyn, associates with macropinosomes via its N-terminus continuously from their generation at membrane ruffles, through their centripetal trafficking, to fusion with late endosomes and lysosomes. c-Src is rapidly recruited to macropinosomal membranes from the plasma membrane and intracellular organelles.
5. Src signaling regulates the terminal step in cytokinesis called abscission in HeLa cells. Completion of abscission requires Src-mediated tyrosine phosphorylation during early stages of mitosis (before cleavage furrow formation), which is subsequently delivered to the midbody through Rab11-driven vesicle transport. Src kinase activity during the early mitosis mediates ERK activation in late cytokinesis. Recruitment of activated ERK, which is phosphorylated by MEK downstream of Src kinases, to the midbody plays an important role in completion of abscission.
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