【摘要】 为了研究Pin1和Rb蛋白的细胞定位、Pin1调控Rb磷酸化的相关机制,本研究构建了慢病毒载体pLVX-Flag-HA-Pin1和Plko.1-shRNA,包装病毒感染人非小细胞肺癌(H1299),免疫印迹检测Pin1蛋白表达水平,采用免疫荧光染色、免疫组化技术研究蛋白的定位.结果表明,沉默Pin1可降低Rb的磷酸化并抑制细胞的增殖;培养的肿瘤细胞和肿瘤组织中,Pin1可定位到细胞质中,而胞质定位的Pin1也可增加Rb的磷酸化.更多还原

【Abstract】 In order to study the intracellular localization of Pin1 and Rb protein and the mechanisms of Pin1 regulated Rb phosphorylation,lentivirus vector pLVX-Flag-HA-Pin1 and Plko.1-shRNA were constructed.After lentiviral packaging and infection,Pin1 protein level were detected by western blot in human non-small cell 1ung carcinoma cell(H1299).Pin1 and Rb protein localization were studied by immunofluorescence staining and immunohistochemistry.The data demonstrated that knockdown of Pin1 result in a decrease of Rb phosphorylation and cellular viability.Pin1 can localize to cytoplasm in cultured tumor cells and tumor tissues,while cytoplasmic Pin1 can increase the phosphorylation of Rb.This is a novel mechanism for Pin1 in regulating Rb function.更多还原