PKC信号调控负性共刺激分子B7-H4在结直肠癌异常表达的作用机制

B7-H4 is a new member of the B7 family, plays an pivotal role in tumor immunologic escape by inhibiting the effects of T cells and promoting the growth and metastasis of tumor cells. However, the regulation mechanism of it remains unknown. We found the coexpression of B7-H4 and PKC-δin colorectal cancer cells was positive related through correlative statistic analysis, and TPA could promote the expression of B7-H4 in colorectal cancer cells. So we hypothesized that PKC passway might play a critical role in the regulation of B7-H4 in tumor. To find the transcription factors(TF) that could bind with B7-H4 promotor, we want to amplify the gene promoter through PCR method, and confirm their binding by Chromatin immunoprecipitation (ChIP) and electrophoretic mobility shift assay(EMSA).And The TF recombinant vectors were transfected into colorectal cancer cells to investigate the reflections of the cells, we also use the mice model to confirm the functions of the TF. In a word, we explored the regulation of B7-H4 gene in this study, which may have a great significance for the function research of B7-H4 gene and set up a new approach for cancer therapy.

B7-H4可负性调控T细胞免疫应答，并促进肿瘤细胞增殖与转移，已成为目前研究热点，但其在肿瘤组织异常表达的调控机制尚未可知。我们发现B7-H4与蛋白激酶C-δ在结直肠癌中存在异常表达且显著正相关，进一步研究发现PKC激活剂TPA可显著促进结直肠癌细胞中B7-H4的表达，据此推测PKC通路可能在肿瘤B7-H4表达调控中发挥了重要作用。本项目拟通过双荧光素酶检测系统、染色质免疫共沉淀和凝胶迁移率实验结合生物信息学分析，借助B7-H4 promotor/PGL3 Basic重组表达载体这一工具，筛选出与B7-H4 promotor结合的转录因子，并确定与之作用的特异性启动子区；同时构建过表达及缺失表达候选转录因子的结直肠癌细胞株，通过体内外实验验证该因子与PKC信号的关联及对肿瘤细胞B7-H4表达的影响。初步阐明B7-H4在结直肠癌异常表达的调控机制，为我们找寻肿瘤治疗的新靶点提供基础。