Follicle stimulating hormone-regulated expression of serum/glucocorticoid-inducible kinase in rat ovarian granulosa cells: a functional role for the Sp1 family in promoter activity.
Follicle stimulating hormone-regulated expression of serum/glucocorticoid-inducible kinase in rat ovarian granulosa cells: a functional role for the Sp1 family in promoter activity.
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DOI：
10.1210/mend.11.13.0033
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发表时间：
1997-12
影响因子：
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通讯作者：
Tamara Alliston;A. C. Maiyar;Patricia Buse;Gary L. Firestone;J. S. Richards
中科院分区：
文献类型：
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作者：
Tamara Alliston;A. C. Maiyar;Patricia Buse;Gary L. Firestone;J. S. Richards
最近，已经鉴定了一个新的丝氨酸/苏氨酸蛋白激酶家族。这些转录诱导，立即早期基因编码血清/糖皮质激素诱导蛋白激酶，sgk。通过原位杂交，我们发现大鼠卵巢中sgk的表达选择性地定位于颗粒细胞。在培养中，FSH或毛喉素，蛋白激酶A（PKA）途径的激活剂，迅速（2小时）和短暂增加未分化颗粒细胞中的sgk mRNA水平。Sgk mRNA表现出双相表达模式，在48小时观察到最大水平的FSH/毛喉素作为颗粒细胞分化为排卵前表型。使用sgk启动子-报告基因构建体（-4.0 kb至-35 bp）的缺失分析鉴定了在未分化和分化的颗粒细胞中介导FSH和毛喉素应答转录的-63和-43 bp之间的区域。该富含G/C的区域1）赋予最小-35 sgk启动子氯霉素乙酰转移酶报告基因构建体基础和诱导转录，2）特异性结合颗粒细胞提取物中存在的Sp1和Sp3，以及3）结合重组Sp1。在该区域中的2 bp的突变不仅阻止了Sp1和Sp3结合，而且还消除了使用野生型构建体时观察到的PKA介导的反式激活。Sp1和Sp3的DNA结合活性和蛋白水平在sgk诱导过程中没有显着变化。总的来说，这些数据表明，Sp1/Sp3的sgk启动子的反式激活可能涉及调节，磷酸化依赖性与其他因素的相互作用。因此，与颗粒细胞从增殖到分化的进展相关的sgk的新的双相诱导似乎涉及FSH、PKA和转录因子（包括Sp1和Sp3）的顺序协调作用。
Recently, a family of novel, serine/threonine protein kinases has been identified. One of these transcriptionally inducible, immediate-early genes encodes serum/glucocorticoid inducible-protein kinase, sgk. By in situ hybridization, we show that sgk expression in the rat ovary is selectively localized to granulosa cells. In culture, FSH or forskolin, activators of the protein kinase A (PKA) pathway, rapidly (2 h) and transiently increased sgk mRNA levels in undifferentiated granulosa cells. Sgk mRNA exhibited a biphasic expression pattern, with maximal levels observed at 48 h of FSH/forskolin as granulosa cells differentiate to the preovulatory phenotype. Deletion analyses using sgk promoter-reporter constructs (-4.0 kb to -35 bp) identified a region between -63 and -43 bp that mediated FSH and forskolin-responsive transcription in undifferentiated and differentiated granulosa cells. This G/C-rich region 1) conferred both basal and inducible transcription to the minimal -35 sgk promoter chloramphenicol acetyltransferase reporter construct, 2) specifically bound Sp1 and Sp3 present in granulosa cell extracts, and 3) bound recombinant Sp1. Mutation of 2 bp in this region not only prevented Sp1 and Sp3 binding, but also abolished the PKA-mediated transactivation observed when using the wild type construct. Sp1 and Sp3 DNA-binding activity and protein levels did not change significantly during sgk induction. Collectively, these data indicate that Sp1/Sp3 transactivation of the sgk promoter likely involves regulated, phosphorylation-dependent interaction with other factors. Thus the novel, biphasic induction of sgk that correlates with granulosa cell progression from proliferation to differentiation appears to involve sequential, coordinated actions of FSH, PKA, and transcription factors, including Sp1 and Sp3.