脂筏微域内CD59趋引CBP蛋白介导T细胞下游信号的转导研究  

CD59 recruits Cbp into lipid raft to modulate the signal transduction in T lymphocytes

作　　者：丛蓓蓓[1] 高美华[1] 王冰[1] 王丽娜[1] 邵志伟[2] 

机构地区：[1]青岛大学医学院免疫教研室,266071 [2]青岛大学医学院神经外科,266071

出　　处：《免疫学杂志》2015年第1期1-6,共6页Immunological Journal

摘　　要：目的探讨Cbp(Csk-binding protein)在Jurkat细胞中的过表达对其生物学功能的作用以及GPI锚定蛋白CD59通过Cbp对细胞的效应机制。方法构建Cbp-EGFP慢病毒载体,转染建立稳定的过表达Cbp-EGFP的Jurkat细胞株。采用免疫荧光技术,通过共聚焦显微镜观察CD59、Cbp的表达及定位关系;检测CD59抗体刺激前后细胞的增殖效率及细胞凋亡情况;Western blot检测Jurkat细胞中信号转导通路相关蛋白分子的变化。结果共聚焦显微镜下可见CD59、Cbp定位于胞膜上,在抗体刺激后,Cbp组与对照组均出现明显的荧光高亮聚集现象。转染Cbp-EGFP后细胞增殖速率明显降低(P<0.05),而中晚期凋亡现象显著高于阴性对照组(P<0.05)。CD59单抗作用前后相比,酪氨酸激酶蛋白(Fyn、Lck、ZAP-70)的表达量均有所下降,而Csk的表达量增加。结论 Cbp过表达能有效抑制Jurkat细胞的增殖,GPI锚固蛋白CD59可通过Cbp分子调控细胞信号转导通路中重要蛋白酪氨酸激酶的表达,进而影响Jurkat细胞的生物学活性。This study designed to investigate the effects of over-expressed Cbp （Csk-binding protein） on the biological functions of T lymphocytes, as well as the effector mechanism about GPI-anchored protein CD59 acting on the Jurkat cells by Cbp. Firstly, a Cbp-EGFP lentiviral vector was constructed for establishing stable cell lines high expressing Cbp-EGFP through transfection. Then, the expression and location of CD59 and Cbp were observed by immunofluorescence cytochemistry; the proliferation and apoptosis of the cells were measured before and after CD59 antibody stimulation; and the changes of proteins in Jurkat cells were detected by Western blot. We found that CD59 and Cbp were mainly distributed in cell membrane, and appeared punctate aggregation phenomenon after stimulation by CD59 antibody. After transfection of Cbp-EGFP, cell proliferation rate was significantly decreased （P〈 0.05）, and the apoptosis phenomenon was significantly obvious than that in control group （P〈 0.05）. Furthermore, CD59 monoclonal antibody stimulation could decrease the protein tyrosine kinase （Fyn, Lck, ZAP- 70） expression, but increase the expression of Csk. Together all, the over expression of Cbp gene can effectively inhibit the proliferation of Jurkat cells; GPI-anchored protein CD59 can regulate the expression of important tyrosine kinase proteins in cell signal transduction pathway through Cbp molecular, then affect the biological activities of Jurkat cells.

关 键 词：CD59 CBP T淋巴细胞 信号转导 

分 类 号：R392[医药卫生—免疫学]