Huang N, Miller W L
Department of Pediatrics, University of California, San Francisco, San Francisco, California 94143-0978, USA.
J Biol Chem. 2000 Jan 28;275(4):2852-8. doi: 10.1074/jbc.275.4.2852.
The cholesterol side-chain cleavage enzyme, cytochrome P450scc, initiates the biosynthesis of all steroid hormones. Adrenal and gonadal strategies for P450scc gene transcription are essentially identical and depend on the orphan nuclear receptor steroidogenic factor-1, but the placental strategy for transcription of P450scc employs cis-acting elements different from those used in the adrenal strategy and is independent of steroidogenic factor-1. Because placental expression of P450scc is required for human pregnancy, we sought factors that bind to the -155/-131 region of the human P450scc promoter, which participates in its placental but not adrenal or gonadal transcription. A yeast one-hybrid screen of 2.4 x 10(6) cDNA clones from human placental JEG-3 cells yielded two unique clones; one is the previously described transcription factor LBP-1b, which is induced by HIV, type I infection of lymphocytes, and the other is a new factor, termed LBP-9, that shares 83% amino acid sequence identity with LBP-1b. When expressed in transfected yeast, both factors bound specifically to the -155/-131 DNA; antisera to LBP proteins supershifted the LBP-9.DNA complex and inhibited formation of the LBP-1b.DNA complex. Reverse transcriptase-polymerase chain reaction detected LBP-1b in human placental JEG-3, adrenal NCI-H295A, liver HepG2, cervical HeLa, and monkey kidney COS-1 cells, but LBP-9 was detected only in JEG-3 cells. When the -155/-131 fragment was linked to a minimal promoter, co-expression of LBP-1b increased transcription 21-fold in a dose-dependent fashion, but addition of LBP-9 suppressed the stimulatory effect of LBP-1b. The roles of LBP transcription factors in normal human physiology have been unclear. Their modulation of placental but not adrenal P450scc transcription underscores the distinctiveness of placental strategies for steroidogenic enzyme gene transcription.
胆固醇侧链裂解酶，即细胞色素P450scc，启动了所有甾体激素的生物合成。肾上腺和性腺中P450scc基因转录的调控机制基本相同，均依赖于孤儿核受体类固醇生成因子-1，但胎盘组织中P450scc基因转录的调控机制则采用了与肾上腺不同的顺式作用元件，且不依赖于类固醇生成因子-1。由于人类妊娠需要胎盘组织表达P450scc，我们寻找了能与人类P450scc启动子-155/-131区域结合的因子，该区域参与胎盘组织而非肾上腺或性腺组织中的转录过程。用人胎盘JEG-3细胞的2.4×10⁶个cDNA克隆进行酵母单杂交筛选，得到了两个独特的克隆；一个是先前描述的转录因子LBP-1b，它可由HIV-1感染淋巴细胞诱导产生，另一个是新的因子，称为LBP-9，其与LBP-1b的氨基酸序列同一性为83%。当在转染的酵母中表达时，这两个因子均能特异性地结合-155/-131 DNA；针对LBP蛋白的抗血清使LBP-9.DNA复合物出现超迁移，并抑制LBP-1b.DNA复合物的形成。逆转录-聚合酶链反应在人胎盘JEG-3、肾上腺NCI-H295A、肝脏HepG2、宫颈HeLa和猴肾COS-1细胞中检测到了LBP-1b，但仅在JEG-3细胞中检测到了LBP-9。当-155/-131片段与最小启动子相连时，LBP-1b共表达可使转录以剂量依赖的方式增加21倍，但加入LBP-9可抑制LBP-1b的刺激作用。LBP转录因子在正常人体生理学中的作用尚不清楚。它们对胎盘组织而非肾上腺组织中P450scc转录的调节突出了胎盘组织中类固醇生成酶基因转录调控机制的独特性。
