摘要
目的 :初步分析小鼠α2 ( )胶原基因上游 5′- U TR近端 80 0 bp(- 780～ + 5 4bp)序列的 DNA结合蛋白。 方法 :PCR扩增小鼠α2 ( )胶原基因 - 2 5 8～ + 5 4bp,- 5 19～ - 2 48bp,- 741～ - 5 0 1bp片段 ,Dig- d U TP末端标记 PCR产物 ,与经 SDS- PAGE并转印至膜上的胶原产生细胞的核抽提物进行 DNA -蛋白质结合反应 ,以抗 Dig抗体检测结合反应信号。 结果 : 型胶原基因上游 5′- U TR近端 80 0 bp序列中存在至少 3个不同的核转录因子结合位点 ,相识别 (consensus)的核蛋白因子相对分子质量分别为 12万 ,5万 ,2万。TGF- β1可增强细胞中相对分子质量为 12万的核蛋白因子的表达。结论 :小鼠 α2( )胶原基因上游 - 780～ + 5 4bp这 80 0 bp片段中存在与不同的核蛋白因子结合的序列 ,可能与 Sp- 1有关。TGF- β1通过增强相对分子质量为 12万核蛋白的表达而促进该蛋白与靶调控序列的结合。
Objective:To probe the consensus DNA binding proteins of proximal 800 bp(-780 to +54 bp) fragment in mouse α2(Ⅰ) procollagen gene. Methods:Three fragments(-258 to +54 bp, -519 to -248 bp, -741 to -501 bp) were amplified by PCR, end labeled with Dig dUTP and hybridized with nuclear extracts from collagen producing cells which were electrophoresed and blotted onto nitrocellulose membrane . Results: The DNA protein binding signals were captured by immunological detection. At least 3 different binding sites for nuclear transcription factor existed in proximal 800 bp fragment in mouse α2(Ⅰ) procollagen gene. The molecular weight of consensus nuclear factors were 12×10 4, 5×10 4, 2×10 4 respectively. The expression of 12×10 4 nuclear protein was enhanced by TGF β1 . Conclusion: Sp 1 might be among the several consensus nuclear transcription factors of -780 to +54 bp sequence in mouse α2(Ⅰ) procollagen gene. The positive effect of TGF β1 on procollagen gene might be mediated through enhanced expression of the nuclear factor with molecular weight 12×10 4. [
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2000年第8期704-707,共4页
Academic Journal of Second Military Medical University
基金
国家自然科学基金!资助项目 (395 0 0 0 6 8
39870 30 1)
长征医院"208"计划资助
