Luteinizing hormone beta promoter stimulation by adenylyl cyclase and cooperation with gonadotropin-releasing hormone 1 in transgenic mice and LBetaT2 cells
Luteinizing hormone beta promoter stimulation by adenylyl cyclase and cooperation with gonadotropin-releasing hormone 1 in transgenic mice and LBetaT2 cells
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DOI：
10.1095/biolreprod.107.064139
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发表时间：
2007-12-01
影响因子：
3.6
通讯作者：
Shupnik, Margaret A.
中科院分区：
文献类型：
--
作者：
Ferris, Heather A.;Walsh, Heidi E.;Shupnik, Margaret A.
下丘脑促性腺激素释放激素1 （GnRH1）刺激大鼠促黄体生成素β (Lhb)基因转录，这种反应可能受到cAMP等其他信号通路的调节。在这里，我们描述了cAMP单独或与GnRH1一起刺激小鼠垂体和克隆性腺促细胞中Lhb基因转录的能力。cAMP和垂体腺苷酸环化酶激活肽均能增加表达大鼠lhb -荧光素酶转基因小鼠垂体中GnRH1对荧光素酶活性的刺激，提示cAMP和GnRH1途径在体内相互作用。lhb -荧光素酶转基因cAMP对雌性发情期和发情期的刺激相似，但GnRH1对发情期的刺激更大。两种治疗方法的加性效应均见于孕前期和发情期。升高的细胞内cAMP刺激LbetaT2克隆促性腺激素细胞中Lhb启动子活性，单独或与GnRH1联合。在LbetaT2细胞中，cAMP对Lhb启动子的刺激通过抑制蛋白激酶A （PKA）而被消除；PKA或蛋白激酶C抑制剂均可部分抑制GnRH1的刺激。cAMP刺激只需要启动子的近端gnrh1响应区域，而3' NR5A1位点的突变会减弱这种反应。cAMP和GnRH1对内源性Lhb基因初级mRNA转录物的调控与Lhb-荧光素酶转基因或转染启动子的结果相关。添加或不添加forskolin时，GnRH1增加了EGR1对内源性启动子的占用，但单独使用forskolin影响不大。因此，cAMP对Lhb启动子活性的刺激和GnRH1刺激的增强，通过pka依赖的机制发生在多种独立于类固醇状态的生理状态中。
Rat luteinizing hormone beta (Lhb) gene transcription is stimulated by hypothalamic gonadotropin-releasing hormone 1 (GnRH1), and this response may be modulated by other signaling pathways such as cAMP. Here we characterize the ability of cAMP, alone or with GnRH1, to stimulate Lhb gene transcription in mouse pituitary and clonal gonadotroph cells. Both cAMP and pituitary adenylyl cyclase-activating peptide increase GnRH1 stimulation of luciferase activity in pituitaries of mice expressing the rat Lhb-luciferase transgene, suggesting cAMP and GnRH1 pathways interact in vivo. cAMP stimulation of the Lhb-luciferase transgene was similar between females in metestrus and proestrus, but GnRH1 stimulation was greater at proestrus. Additive effects with combined treatments were observed at metestrus and proestrus. Elevated intracellular cAMP stimulated Lhb promoter activity in LbetaT2 clonal gonadotroph cells, alone and with GnRH1. In LbetaT2 cells, cAMP stimulation of the Lhb promoter was eliminated by inhibition of protein kinase A (PKA); GnRH1 stimulation was partially suppressed by either PKA or protein kinase C inhibitors. Only the proximal GnRH1-responsive region of the promoter was required for cAMP stimulation, and mutation of the 3' NR5A1 site diminished the response. Regulation of primary mRNA transcripts from the endogenous Lhb gene by cAMP and GnRH1 correlated with results from the Lhb-luciferase transgene or transfected promoter. Occupancy of the endogenous promoter by EGR1 was increased by GnRH1 with or without forskolin, but forskolin alone had little effect. Thus, cAMP stimulation of Lhb promoter activity, and enhancement of GnRH1 stimulation, occurs in multiple physiological states independent of steroid status, via a PKA-dependent mechanism.