PTD-OD-HA融合蛋白对K562细胞中Bcr-Abl的抑制作用观察  

Inhibitory effect of PTD-OD-HA fusion protein on Bcr-Abl in K562 cells

作　　者：高淼[1] 黄峥兰[1] 季茂胜[1] 黄世峰[1] 曾建明[1] 李千音[1] 冯文莉[1] 

机构地区：[1]重庆医科大学临床血液学教研室,临床检验诊断学教育部重点实验室,重庆400016

出　　处：《解放军医学杂志》2012年第10期951-954,共4页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金(30670901)~~

摘　　要：目的探讨PTD-OD-HA融合蛋白转导K562细胞的动力学、定位及其与Bcr-Abl定位的关系,以及对Bcr-Abl寡聚化和Bcr-Abl酪氨酸激酶活性的影响。方法采用FITC标记PTD-OD-HA融合蛋白,观察蛋白转导K562细胞的效率与剂量和时间的关系,激光共聚焦显微镜检测融合蛋白在细胞内的定位,免疫共沉淀法检测融合蛋白与Bcr-Abl的相互作用,Western blotting检测Bcr-Abl的磷酸化水平。结果 PTD-OD-HA转导K562细胞呈剂量和时间依赖性。PTD-OD-HA定位于K562细胞胞质,与Bcr-Abl共定位,且能与Bcr-Abl蛋白相互作用,进而干扰Bcr-Abl同源寡聚化,并可降低Bcr-Abl的磷酸化水平。结论在K562细胞中,PTD-OD-HA融合蛋白能有效抑制Bcr-Abl同源寡聚化及磷酸化水平。Objective To study the transduction dynamics, location of PTD-OD-HA fusion protein and its interaction with Bcr-Abl oncoprotein in K562 cell lines, and explore the influence of PTD-OD-HA fusion protein on oligomerization and tyrosine kinase activity of Bcr-Abl. Methods PTD-OD-HA fusion protein was labeled with FITC and co-cultured with K562 cells. The transduction efficiency of labeled PTD-OD-HA at different doses and time intervals was observed under fluorescence microscope. The location oflabeled PTD-OD-HA fusion protein in K562 cells was detected by confocal microscopy. The interaction of PTD-OD-HA fusion protein with Bcr-Abl oncoprotein was confirmed by coimmunoprecipitation. The phosphorylation of Bcr- Abl oncoprotein was detected by Western blotting. Results PTD-OD-HA fusion protein labeled with FITC was transduced into K562 cells in a dose- and time-dependent manner. PTD-OD-HA fusion protein was located in the cytoplasm of K562 cells and was consistent with the location of Bcr-Abl oncoprotein. The interaction of PTD-OD-HA fusion protein with Bcr-Abl oncoprotein was proved in K562 cells. This interaction could interrupt the homologous oligomerization of Bcr-Abl oncoprotein and reduce the phosphorylation of Bcr-Abl oncoprotein. Conclusion PTD-OD-HA fusion protein could be transduced into K562 cells efficiently, inhibiting the oligomerization and reducing the phosphorylation of Bcr-Abl oncoprotein.

关 键 词：白血病 髓样 慢性 蛋白转导域 融合蛋白质类 bcr-abl 

分 类 号：R733.722[医药卫生—肿瘤]