CAMP REGULATES P450SCC GENE-EXPRESSION BY A CYCLOHEXIMIDE-INSENSITIVE MECHANISM IN CULTURED MOUSE LEYDIG MA-10 CELLS
CAMP REGULATES P450SCC GENE-EXPRESSION BY A CYCLOHEXIMIDE-INSENSITIVE MECHANISM IN CULTURED MOUSE LEYDIG MA-10 CELLS
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DOI：
10.1073/pnas.86.20.7775
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发表时间：
1989-10-01
影响因子：
11.1
通讯作者：
VAISSE, C
中科院分区：
文献类型：
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作者：
MELLON, SH;VAISSE, C
小鼠MA-10 Leydig肿瘤细胞合成并分泌孕酮，对人绒毛膜促性腺激素、促黄体激素和cAMP作出反应，但可能不合成雄激素。最大剂量的人绒毛膜促性腺激素，羊促黄体激素，毛喉素，或8-溴腺苷3 ''，5 ''-环磷酸，刺激细胞色素P450 SCC mRNA的积累1.5至3倍，孕酮分泌10至100倍的MA-10细胞。P450 SCC mRNA在2小时后增加，8小时时达到最大，聚合酶运行实验表明这是由于P450 SCC基因转录增加。MA-10细胞是一个同源的群体，因为所有的细胞表达P450 SCC mRNA，对cAMP的反应相同。在环己酰亚胺的存在下，cAMP刺激的P450 SCC mRNA的积累继续。促性腺激素通过协调cAMP诱导的P450 SCC基因转录、mRNA积累和P450 SCC活性增加来刺激睾丸类固醇生成。我们克隆了大鼠P450 c17 cDNA，并通过RNA转移印迹或RNA酶保护试验表明，在对照或cAMP刺激的MA-10细胞中没有检测到P450 c17 mRNA。类似地，HPLC未检测到17 α-羟孕酮或睾酮合成。因此，MA-10细胞，不像未转化的Leydig细胞，不表达可检测量的P450 c17 mRNA或P450 c17活性。
Mouse MA-10 Leydig tumor cells synthesize and secrete progesterone in response to human chorionic gonadotropin, luteinizing hormone, and cAMP but may not synthesize androgens. Maximal doses of human chorionic gonadotropin, ovine luteinizing hormone, forskolin, or 8-bromoadenosine 3'',5''-cyclic monophosphate, stimulated cytochrome P450scc mRNA accumulation 1.5- to 3-fold and progesterone secretion 10- to 100-fold in MA-10 cells. P450scc mRNA increased by 2 hr and was maximal by 8 hr, polymerase run-on experiments showed this was due to increased P450 scc gene transcription. MA-10 cells are a hormonally homogeneous population, as all cells expressed P450scc mRNA and responded to cAMP equally. cAMP-stimulated accumulation of P450scc mRNA continued in the presence of cycloheximide. Gonadotropins stimulated testicular steroidogenesis by coordinate cAMP-induced increases in P450scc gene transcription, mRNA accumulation, and P450scc activity. We cloned rat P450c17 cDNA and showed it detected no P450c17 mRNA in control or cAMP-stimulated MA-10 cells by RNA transfer blots or RNase protection assays. Similarly, HPLC detected no 17.alpha.-hydroxyprogesterone or testosterone synthesis in MA-10 cells. Thus MA-10 cells, unlike untransformed Leydig cells, do not express detectable amounts of P450c17 mRNA or P450c17 activity.