NPRAP蛋白通过调节小GTP酶活性促进肺癌细胞的恶性转化  

NPRAP protein promotes malignant transformation of lung cancer cells by regulating the activity of small GTPases

作　　者：张俊毅[1] 董彩凤[2] 刘晓辉[1] 林璐璐[1] 于佳乐[1] 刘帅莹[1] 

机构地区：[1]赤峰学院医学院病理教研室,附属医院病理科,内蒙古自治区赤峰024000 [2]赤峰学院附属医院呼吸内科,内蒙古自治区赤峰024000

出　　处：《肿瘤》2013年第6期483-489,共7页Tumor

基　　金：国家自然科学基金资助项目(编号:81201844);内蒙古自治区自然科学基金资助项目(编号:2012MS1109)

摘　　要：目的:探讨NPRAP蛋白表达对肺癌细胞生物学行为的影响及其可能的机制。方法:应用质粒转染和RNA干扰的方法分别调节肺癌细胞株SK-MES-1和NCI-H460中NPRAP的表达水平,蛋白质印迹法检测NPRAP蛋白表达情况后,通过MTT、Transwell、免疫荧光、G-LISA比色和Pull-Down方法检测以上肿瘤细胞的增殖、侵袭、形态及小GTP酶(包括RhoA、Cdc42和Rac1)活性的变化。同时,应用Cdc42和Rac1的活性抑制剂ML141抑制小GTP酶活性后,MTT、Transwell和免疫荧光法观察NPRAP对肿瘤细胞生物学行为的影响。结果:肺癌细胞中过表达NPRAP可以降低RhoA活性,升高Cdc42和Rac1活性,并促进肺癌细胞的增殖、侵袭和伪足形成(P<0.05)。相反地,应用小干扰RNA(small interference RNA,siRNA)下调内源性NPRAP的表达后,RhoA活性升高,Cdc42和Rac1活性降低,且肺癌细胞的增殖和侵袭能力下降,伪足数量减少(P<0.05)。此外,应用ML141可以显著减弱NPRAP对肿瘤细胞以上生物学行为的影响(P<0.05)。结论:NPRAP蛋白可能通过改变小GTP酶的活性,导致细胞骨架重新装配,进而提高了肺癌细胞的恶性程度。Objective： To explore the influence of NPRAP （neural plakophilin-related armadillo protein） expression on biological behavior of lung cancer cells and its possible mechanism. Methods： The expression level of NPRAP was regulated by plasmid transfection and RNA interference methods in two lung cancer cell lines SK-MES-1 and NCI-H460, respectively. Western blotting was used to detect the expression of NPRAP in cancer cells. Then, the abilities of proliferation and invasion and the morphology of the cancer cells were detected by MIT, Transwell, G-LISA and Pull-Down methods, respectively. The activities of small GTPases （including RhoA, Cdc42, and Racl） in cancer cells were detected by immunofluorescence assay. At the same time, ML141, an inhibitor of Cdc42 and Racl activities, was used to explore whether the change of the activity of small GTPases was involved in the influence of NPRAP on biological behavior of cancer cells or not. Results： Over-expression of NPRAP decreased RhoA activity and increased Cdc42/Rac1 activity, and also promoted the proliferation and invasion abilities of lung cancer cells （P 〈 0.05）, as well as the pseudopodia formation. Conversely, the down-regulation of endogenous NPRAP expression level induced by siRNA （small interference RNA） could increase RhoA activity and decrease Cdc42/Rac1 activity, and also inhibit the proliferation, invasion and pseudopodia formation of lung cancer cells （P 〈 0.05）. In addition, the influence of NPRAP on biological behavior of cancer cells was weakened by using ML141 （P 〈 0.05）. Conclusion： NPRAP protein probably results in cytoskeleton reassembly by regulating the activity of small GTPases, and then promotes malignant transformation of lung cancer cells.

关 键 词：肺肿瘤 细胞黏附分子 单体GTP结合蛋白质类 基因表达调控 肿瘤 NPRAP 

分 类 号：R734.2[医药卫生—肿瘤]