Activation of cAMP-dependent protein kinase increases the protein level of steroidogenic factor-1
Activation of cAMP-dependent protein kinase increases the protein level of steroidogenic factor-1
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DOI：
10.1210/en.143.1.295
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发表时间：
2002-01-01
期刊：
影响因子：
4.8
通讯作者：
Lund, J
中科院分区：
文献类型：
--
作者：
AEsoy, R;Mellgren, G;Lund, J
孤儿核受体甾体生成因子1 （SF-1）是内分泌器官发生、性别分化和甾体生成的重要调节因子。SF-1是cAMP应答基因的转录调节因子，但cAMP依赖性PKA调节SF-1依赖性转录导致类固醇生成量增加的确切机制尚未确定。本研究利用SF-1全长cDNA与编码绿色荧光蛋白（GFP）的cDNA融合，研究了PKA活化对活细胞中SF-1的影响。GFP-SF-1融合蛋白在致甾体性Y1细胞和非致甾体性COS-1细胞的细胞核中均有定位，野生型SF-1的功能特性保持不变。当PKA催化亚基与GFP-SF-1共表达时，我们观察到荧光发射明显升高。Western blot分析证实了这些发现，表明PKA刺激增加了SF-1蛋白水平。pka诱导的SF-1蛋白表达不伴有SF-1 mRNA水平的升高。然而，脉冲追踪研究显示，响应PKA的激活，SF-1的降解率降低，这表明PKA通过增加SF-1蛋白的稳定性来提高SF-1的水平。
The orphan nuclear receptor steroidogenic factor 1 (SF-1) is an essential regulator of endocrine organogenesis, sexual differentiation, and steroidogenisis. SF-1 is a transcriptional regulator of cAMP responsive genes, but the exact mechanisms by which cAMP-dependent PKA modulates SF-1 dependent transcription leading to increased steroidogenic output have not been determined. In this report the effects of PKA activation on SF-1 in living cells have been examined by the use of full-length SF-1 cDNA fused to the cDNA encoding green fluorescent protein (GFP). The GFP-SF-1 fusion protein localized to the nucleus of both steroidogenic Y1 cells and non-steroidogenic COS-1 cells, and the functional properties of wild-type SF-1 were conserved. When the catalytic subunit of PKA was coexpressed with GFP-SF-1, we observed that the fluorescence emission was markedly elevated. These findings were confirmed by Western blot analysis, showing that stimulation of PKA increased SF-1 protein levels. The PKA-induced expression of SF-1 protein was not accompanied by an increase in SF-1 mRNA levels. However, pulse-chase studies showed a decrease in SF-1 degradation rate in response to activation of PKA, indicating that PKA elevates the level of SF-1 by increasing the stability of SF-1 protein.