| Budget Amount *help |
¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000)
Fiscal Year 2010: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000)
Fiscal Year 2009: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2008: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
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| Research Abstract |
V-1 is 13 kDa protein with four ankyrin-repeats for specific molecular interaction with CapZ, an actin capping protein, which was originally identified by us as a developmentally regulated protein in murine postnatal brains. Our recent in vitro and in vivo studies have demonstrated that V-1 can positively control gene transcription of tyrosine hydroxylase (TH), the rate-limiting enzyme for catecholamine biosynthesis, thereby promoting catecholamine production in catecholaminergic cells. In the present study, we tried to elucidate the molecular mechanism underlying control of TH gene expression by V-1 in PC12D cells, a rat adrenal pheochromocytoma cell line endogenously expressing TH protein, using siRNAs for V-1, CapZ and pref-1/DLK1, because the pref-1/DLK1 gene expression is observed to be up-regulated by V-1 in MC3T3-G2/PA6 cells, a mouse preadipocyte cell line. Knockdown of V-1 and CapZ with each siRNA for the both genes resulted in a decrease in TH protein level, accompanied by down-regulation of the expression of pref-1/DLK1 protein, whereas actin protein level showed no changes, as assayed by Western blot analyses. Consistently, it was shown that knockdown of pref-1/DLK1 caused decreased expression of TH protein. Moreover, TH gene transcription is decreased by knockdown of V-1, CapZ and pref-1/DLK1 with each of siRNAs for the three genes, when determined by reporter gene assay. These findings suggest that V-1 may control TH gene expression via pref-1/DLK1 protein in co-operation with CapZ, the actin cytoskeletal regulator, in the CNS catecholaminergic cells.
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