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Nitric oxide inhibits Oct-1 DNA binding activity in cultured vascular smooth muscle cells.

机译:一氧化氮抑制培养的血管平滑肌细胞中的Oct-1 DNA结合活性。

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摘要

Since Oct-1 is a ubiquitous DNA binding protein shown to play an important role in regulating cell proliferation and possess structural characteristics consistent with a nitric oxide (NO) target, we studied NO regulation of the DNA binding activity of Oct-1 in the A7R5 vascular smooth muscle cell (VSMC) line. Two NO donors, sodium nitroprusside (SNP) and S-nitroso-N-acetyl-penicillamine (SNAP) were directly added to the nuclear extract-oligonucleotide reaction mixture, respectively and the effect on Oct-1 DNA binding activity was evaluated by gel shift assay. Both NO donors (0.01-1 mM) inhibited the DNA binding activity of Oct-1. This inhibitory effect was not attenuated by dithiothreitol (DTT) (1 mM) while in contrast, DTT did antagonize the effect of diamide on Oct-1 DNA binding activity. The NO effect on Oct-1 has some specificity; as the NO donors had no effect on myc DNA binding activity. The inhibitory effect of NO donors was reproduced in A7R5 cells, without affecting their viability. These findings provide the first evidence that NO inhibits the DNA binding activity of Oct-1, probably through a cGMP independent mechanism and suggests that NO may inhibit mitogenesis in part through an effect on Oct-1 DNA binding activity in VSMCs.
机译:由于Oct-1是普遍存在的DNA结合蛋白,在调节细胞增殖中起着重要作用,并具有与一氧化氮(NO)靶标一致的结构特征,因此我们研究了NO对A7R5中Oct-1 DNA结合活性的调控血管平滑肌细胞(VSMC)线。将两个NO供体硝普钠(SNP)和S-亚硝基-N-乙酰基-青霉胺(SNAP)直接添加到核提取物-寡核苷酸反应混合物中,并通过凝胶位移评估对Oct-1 DNA结合活性的影响分析。两种NO供体(0.01-1 mM)均抑制Oct-1的DNA结合活性。二硫苏糖醇(DTT)(1 mM)不能减弱这种抑制作用,相反,DTT确实拮抗了二酰胺对Oct-1 DNA结合活性的作用。 NO对Oct-1的影响具有某些特异性。因为NO供体对myc DNA结合活性没有影响。 NO供体的抑制作用在A7R5细胞中复制,而不会影响其生存能力。这些发现提供了第一个证据,即NO可能通过cGMP独立机制抑制了Oct-1的DNA结合活性,并表明NO可能部分地通过影响VSMC中Oct-1 DNA的结合活性来抑制有丝分裂。

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