Mukai K, Mitani F, Shimada H, Ishimura Y
Department of Biochemistry, School of Medicine, Keio University, Tokyo, Japan.
Mol Cell Biol. 1995 Nov;15(11):6003-12. doi: 10.1128/MCB.15.11.6003.
The CYP11B1 gene, which encodes steroid 11 beta-monooxygenase, which is responsible for the synthesis of cortisol and corticosterone, the major glucocorticoids in mammals, is expressed specifically in the zona fasciculata of the adrenal cortex. We have analyzed the promoter region of the rat CYP11B1 gene by using a transient-expression system with adrenocortical Y1 cells and have identified a positive regulatory region. The region contained two adjacent sites for the binding of Y1-cell nuclear proteins: the binding site for an AP-1 transcription factor composed of JunD and a Fos-related protein, and the site for Ad4-binding protein (Ad4BP). The binding of the AP-1 factor to the regulatory region had a suppressive effect on that of Ad4BP in the nuclear extracts. Mutational analyses revealed that the transcriptional activation of the CYP11B1 gene promoter in Y1 cells was attributable to the AP-1 site but not to the Ad4 site. Subsequently, nuclear extracts of the zona fasciculata cells from the rat adrenal cortex were found to contain both AP-1 factor and Ad4BP, whose binding properties to the regulatory region were almost identical to those of the two factors in the Y1-cell nuclear extracts. Moreover, immunohistochemical analyses of rat adrenal cortices showed that the AP-1 factor was present in the nuclei of CYP11B1-expressing cells in the zona fasciculata but not in the nuclei of cells in the other zones. From these results, we propose that the AP-1 transcription factor found in this study plays an important role in the zone-specific expression of the CYP11B1 gene in rat adrenal cortex.
CYP11B1基因编码类固醇11β-单加氧酶，该酶负责合成皮质醇和皮质酮，这两种物质是哺乳动物体内主要的糖皮质激素，该基因在肾上腺皮质束状带中特异性表达。我们利用肾上腺皮质Y1细胞的瞬时表达系统分析了大鼠CYP11B1基因的启动子区域，并确定了一个正调控区域。该区域包含两个相邻的Y1细胞核蛋白结合位点：一个由JunD和一个Fos相关蛋白组成的AP-1转录因子结合位点，以及Ad4结合蛋白（Ad4BP）结合位点。在核提取物中，AP-1因子与调控区域的结合对Ad4BP的结合有抑制作用。突变分析表明，Y1细胞中CYP11B1基因启动子的转录激活归因于AP-1位点而非Ad4位点。随后发现，大鼠肾上腺皮质束状带细胞的核提取物中同时含有AP-1因子和Ad4BP，它们与调控区域的结合特性与Y1细胞核提取物中的这两种因子几乎相同。此外，对大鼠肾上腺皮质的免疫组织化学分析表明，AP-1因子存在于束状带中表达CYP11B1的细胞的细胞核中，而不存在于其他区域细胞的细胞核中。根据这些结果，我们提出本研究中发现的AP-1转录因子在大鼠肾上腺皮质CYP11B1基因的区域特异性表达中起重要作用。
