Keren-Tal I, Dantes A, Plehn-Dujowich D, Amsterdam A
Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel.
Mol Cell Endocrinol. 1997 Mar 14;127(1):49-57. doi: 10.1016/s0303-7207(96)03989-5.
Adrenal binding protein 4 (Ad4BP) known also as steroidogenic factor 1 (SF-1) is a cell specific transcription factor regulating all steroidogenic P450 genes and is present exclusively in steroidogenic tissues. In this study, we examined whether Ad4BP expression is affected by oncogene-induced cell transformation. Using a gel shift assay we report here that nuclear extracts of steroidogenic granulosa cell lines, transformed by SV40 DNA and the Ha-ras oncogene show specific binding activity towards an Ad4 recognition sequence oligonucleotide. In contrast, nuclear extracts obtained from granulosa cells transformed with SV40 alone, which lost their steroidogenic activity, did not exhibit any binding to the Ad4 oligonucleotide. Using a specific antibody to Ad4BP, it was demonstrated that only the steroidogenic cell lines, i.e. transfected with SV40 + Ha-ras, expressed significant amount of the protein. No binding activity to the Ad4 oligonucleotide was evident in fibroblasts transformed with the same oncogenes (SV40 + Ha-ras). Steroidogenic activity in SV40 + Ha-ras transformed granulosa cells was markedly elevated following forskolin or follice stimulating factor (FSH) and further augmented by incubation of the cells with dexamethasone. However, no change in Ad4BP expression and binding activity was observed following such stimulations. It is suggested that Ha-ras expression in SV40 transformed granulosa cells can play an important role in restoring Ad4BP expression and activity, which are required for their steroidogenic function. Thus, expression of Ad4BP is essential for steroidogenesis both in primary and in oncogene transformed granulosa cells.
肾上腺结合蛋白4（Ad4BP），也被称为类固醇生成因子1（SF-1），是一种细胞特异性转录因子，可调节所有类固醇生成的细胞色素P450基因，且仅存在于类固醇生成组织中。在本研究中，我们检测了Ad4BP表达是否受癌基因诱导的细胞转化影响。通过凝胶迁移实验，我们在此报告，经SV40 DNA和Ha-ras癌基因转化的类固醇生成颗粒细胞系的核提取物，对Ad4识别序列寡核苷酸显示出特异性结合活性。相反，仅用SV40转化的颗粒细胞，其类固醇生成活性丧失，其核提取物对Ad4寡核苷酸未表现出任何结合。使用针对Ad4BP的特异性抗体，证明只有类固醇生成细胞系，即转染了SV40 + Ha-ras的细胞系，表达大量该蛋白。用相同癌基因（SV40 + Ha-ras）转化的成纤维细胞中，未观察到对Ad4寡核苷酸的结合活性。用福斯可林或促卵泡刺激素（FSH）处理后，SV40 + Ha-ras转化的颗粒细胞中的类固醇生成活性显著升高，用 dexamethasone孵育细胞后进一步增强。然而，此类刺激后未观察到Ad4BP表达和结合活性的变化。提示SV40转化的颗粒细胞中Ha-ras的表达，在恢复其类固醇生成功能所需的Ad4BP表达和活性方面可发挥重要作用。因此，Ad4BP的表达对于原代和癌基因转化的颗粒细胞中的类固醇生成均至关重要。
