Jimenez Patricia, Saner Karla, Mayhew Bobbie, Rainey William E
Department of Obstetrics and Gynecology, Division of Reproductive Endocrinology and Infertility, University of Texas Southwestern Medical Center, Dallas, Texas 75235-9032, USA.
Endocrinology. 2003 Oct;144(10):4285-8. doi: 10.1210/en.2003-0472. Epub 2003 Jul 31.
GATA-6 and GATA-4 are members of a family of transcription factors (GATA 1-6) that share conserved zinc-finger DNA binding domains. Using semiquantitative RT-PCR, we found that the human adrenal expresses mRNA for GATA-6 but not GATA-4. A recent study showed GATA-6 expression in the adrenal reticularis, the source of adrenal androgens. To investigate the role of GATA-6 in regulation of adrenal cell steroidogenesis, luciferase reporter constructs containing the 5'-flanking DNA from steroidogenic acute regulatory protein, cholesterol side-chain cleavage (CYP11A), 17alpha-hydroxylase (CYP17), and dehydroepiandrosterone-sulfotransferase (SULT2A1) were cotransfected with an expression vector containing GATA-6 into adrenal NCI-H295R cells and nonsteroidogenic HEK293 cells. All promoter/reporter constructs were increased by GATA-6 in the adrenal model. However, in the HEK293 cells only SULT2A1 reporter activity was increased by GATA-6. One key difference between H295R and HEK293 cell lines is the differential expression of steroidogenic factor 1 (SF1). Transfection of HEK293 cells with both GATA-6 and SF1 significantly increased transcriptional activation of all reporter constructs above the effect of GATA-6 or SF1 alone. To determine whether the action of GATA-6 required SF1, we transfected HEK293 cells with each promoter construct plus and minus GATA-6, SF1, and/or the orphan nuclear repressor DAX1. DAX1 opposed SF1-activated transcription of many genes and abolished the GATA-6/SF1 ability to increase reporter activity. These results suggest that the adrenal uses GATA-6 to enhance transcription of steroid-metabolizing enzymes needed to produce dehydroepiandrosterone sulfate. Additionally, GATA-6 works in synergy with SF1 to maximally increase expression of enzymes needed to produce adrenal androgens.
GATA-6和GATA-4是转录因子家族（GATA 1-6）的成员，它们共享保守的锌指DNA结合结构域。通过半定量逆转录聚合酶链反应（RT-PCR），我们发现人类肾上腺表达GATA-6的信使核糖核酸（mRNA），但不表达GATA-4的mRNA。最近的一项研究表明，GATA-6在肾上腺雄激素来源的肾上腺网状带中表达。为了研究GATA-6在调节肾上腺细胞类固醇生成中的作用，将含有类固醇生成急性调节蛋白、胆固醇侧链裂解酶（CYP11A）、17α-羟化酶（CYP17）和硫酸脱氢表雄酮硫酸转移酶（SULT2A1）5'-侧翼DNA的荧光素酶报告基因构建体与含有GATA-6的表达载体共转染到肾上腺NCI-H295R细胞和非类固醇生成的HEK293细胞中。在肾上腺模型中，所有启动子/报告基因构建体均被GATA-6增强。然而，在HEK293细胞中，只有SULT2A1报告基因活性被GATA-6增强。H295R细胞系和HEK293细胞系之间的一个关键差异是类固醇生成因子1（SF1）的差异表达。用GATA-6和SF1共转染HEK293细胞，显著增强了所有报告基因构建体的转录激活，其效果超过单独使用GATA-6或SF1的作用。为了确定GATA-6的作用是否需要SF1，我们用每个启动子构建体分别与GATA-6、SF1和/或孤儿核抑制因子DAX1一起或不一起转染HEK293细胞。DAX1对抗许多基因的SF1激活转录，并消除了GATA-6/SF1增强报告基因活性的能力。这些结果表明，肾上腺利用GATA-6来增强产生硫酸脱氢表雄酮所需的类固醇代谢酶的转录。此外，GATA-6与SF1协同作用，最大程度地增加产生肾上腺雄激素所需酶的表达。
