ptd-od-ha融合蛋白对k562细胞中bcr-abl的抑制作用观察.pdf

Med J Chin PLA, Vol. 37, No. 10, October 1, 2012 　781 PTD-OD-HA融合蛋白对K562细胞中Bcr-Abl 的抑制作用观察 高淼，黄峥兰，季茂胜，黄世峰，曾建明，李千音，冯文莉 [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。 [关键词]　白血病，髓样，慢性；蛋白转导域；融合蛋白质类，bcr-abl [中图分类号]　 R733.722 [文献标志码]　A [文章编号]　0577-7402(2012)10-0781-04 Inhibitory effect of PTD-OD-HA fusion protein on Bcr-Abl in K562 cells * GAO Miao, HUANG Zheng-lan, J I Mao-sheng, HUANG Shi-feng, ZENG Ji an-ming, LI Qian-yin, FENG Wen-li Department of Clinical Hematology, Chongqing Medical University, Chongqing 400016, China * Corresponding author, E-mail: fengwlcqmu@ This work was supported by the National Natural Science Foundation of China [Abstract]　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-culture d with K562 cells. The transduction efficiency of labeled PTD-OD-HA at different doses and time intervals was observed under fluorescence microscope. The location of labeled 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 oncop