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Molecular mechanism of the transcription regulation by the retinoic acid receptor in human salivary gland cell line HSG.

人唾液腺细胞系HSG中视黄酸受体转录调控的分子机制。

基本信息

批准号：
11671850
负责人：
KYAKUMOTO Seiko
金额：
$ 1.86万
依托单位：
Iwate Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671850/
关键词：
retinoic acid retinoic acid receptor coactivator corepressor COUP-TF histone acetyltransferase salivary gland cell line CBP ヒストンアセチル化酵素

项目摘要

Growth of HSG cells is regulated by retinoic acid (RA). Recently, it has been revealed that, in the process of RA signaling, retinoic acid receptors (RAR) interact with coactivators such as CREB-binding protein (CBP) and p160 family member proteins, which facilitates the access of core transcription factors to the DNA.To investigate the relationship of coactivators to the RA signaling in HSG cells, we examined the expression of coactivators. Immunoprecipitation and western blotting revealed the expression of CBP and steroid receptor coactivator 1 (SRC 1), which exhibited the activity of histone acetyltransferase. The overexpression of CBP in HSG cell strongly increased the RA-dependent transcription activation approximately 10-fold. This increase of the transactivation was inhibited by the transfection of the antisense oligonucleotide for CBP.These findings suggest that CBP expressed in HSG cells mediates the growth-regulating transcription activation in concert with RARs.We have previously cloned the DNA fragment of COUP-transcription factor I (COUP-TFI). In this study, the expression of full length COUP-TFI was confirmed by use of RT-PCR and western blotting. To determine the role of COUP-TFI in the RA signaling, the reporter gene analysis was examined. The overexpression of COUP-TFI suppressed the RA-induced transcription activation of the reporter gene. Similar results were shown using a chromatin-integrated stably-transfected reporter gene system. The antisense oligonucleotide for COUP-TFI squelched the RA-dependent growth inhibition which was measured by the [^3H]thymidine incorporation. From these results, COUP-TFI very likely regulates RA-sensitive processes such as proliferation or differentiation of the cells by repressing the RA-induced transactivation.

HSG细胞的生长受视黄酸（RA）的调节。近年来研究发现，在RA信号转导过程中，视黄酸受体（retinoicacidreceptors，RAR）与协同激活因子CREB结合蛋白（CREB bindingprotein，CBP）和p160家族成员蛋白相互作用，促进核心转录因子进入DNA，为探讨协同激活因子与RA信号转导的关系，我们检测了HSG细胞中协同激活因子的表达。免疫沉淀和蛋白质印迹显示CBP和类固醇受体辅激活因子1（SRC 1）的表达，其具有组蛋白乙酰转移酶的活性。CBP在HSG细胞中的过表达强烈增加RA依赖的转录激活约10倍。CBP反义寡核苷酸转染HSG细胞后，CBP的转录激活被抑制，提示CBP与RAR共同介导HSG细胞的生长调节转录激活。在本研究中，全长COUP-TFI的表达通过使用RT-PCR和蛋白质印迹法进行了确认。为了确定COUP-TFI在RA信号传导中的作用，检查了报告基因分析。COUP-TFI的过表达抑制了RA诱导的报告基因的转录激活。使用染色质整合的稳定转染的报告基因系统显示了类似的结果。COUP-TFI的反义寡核苷酸可抑制RA依赖的生长抑制作用，其抑制作用通过[^3H]胸苷掺入来测定。从这些结果来看，COUP-TFI很可能通过抑制RA诱导的反式激活来调节RA敏感的过程，例如细胞的增殖或分化。