摘要
目的:探讨组蛋白去乙酰化酶抑制剂曲古菌素A(TSA)对血管平滑肌细胞(VSMCs)p27kip1表达的影响和调控机制。方法:半定量逆转录聚合酶链反应(RT-PCR)检测p27kip1 mRNA水平,蛋白印迹测定p27kip1和skp2蛋白表达,荧光分光光度法测定20S蛋白酶体活性。结果:100μg/LTSA不影响VSMCs中p27kip1的mRNA水平。100μg/LTSA显著抑制血清诱导的p27kip1蛋白下调,并延长p27kip1蛋白的半衰期。100μg/LTSA抑制血清诱导的skp2表达上调,且skp2表达与相应时点p27kip1蛋白呈负相关。100μg/LTSA对20S蛋白酶体活性物无影响。结论:TSA对VSMCs的p27kip1表达调控不是在转录水平上,而是通过翻译后机制抑制血清诱导VSMCs的p27kip1蛋白降解,其机制可能与TSA抑制泛素连接酶亚单位skp2表达有关。
AIM: To investigate the effect of trichostatin A (TSA) on p27kip1 gene expression in vascular smooth muscle cells. METHODS: Reverse transcription - polymerase chain reaction ( RT - PCR) was used to measure the level of p27kip1 mRNA. The protein levels of p27kip1 and S - phase kinase - associated protein - 2 ( skp2 ) were determined by Western blotting. 20S proteasome activity was quantified by using a fluorogenic proteasome - specific substrate. RESULTS: TSA did not affect mRNA level of p27kip1 in VSMCs, but attenuated serum - induced downregulation of p27kipl through stabilizing p27kip1 turnover. In addition, TSA decreased the expression of skp2, an F- box protein that targets p27kip1 for degradation, but had no effect on proteasome activity. CONCLUSION: TSA regulates p27kip1 expression at the post -translational level in VSMCs.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2008年第7期1313-1316,共4页
Chinese Journal of Pathophysiology