Complex role of the mitochondrial targeting signal in the function of steroidogenic acute regulatory protein revealed by bacterial artificial chromosome transgenesis in vivo.
Complex role of the mitochondrial targeting signal in the function of steroidogenic acute regulatory protein revealed by bacterial artificial chromosome transgenesis in vivo.
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DOI：
10.1210/me.2007-0493
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发表时间：
2008-04
影响因子：
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通讯作者：
G. Sasaki;T. Ishii;P. Jeyasuria;Y. Jo;Assaf Bahat;J. Orly;T. Hasegawa;K. Parker
中科院分区：
文献类型：
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作者：
G. Sasaki;T. Ishii;P. Jeyasuria;Y. Jo;Assaf Bahat;J. Orly;T. Hasegawa;K. Parker
类固醇急性调节蛋白（StAR）通过促进胆固醇向线粒体内膜的传递，刺激肾上腺皮质和性腺中类固醇激素的调节产生。为了探索真正的类固醇生成细胞中StAR功能的关键方面，我们使用转基因小鼠模型来探索体内StAR蛋白的功能。我们首先通过将增强的绿色荧光蛋白靶向肾上腺皮质和性腺的类固醇细胞来验证这种转基因细菌人工染色体重组系统。随后，我们针对野生型StAR （WT-StAR）或缺乏线粒体靶向信号的突变StAR蛋白（N47-StAR）的表达。在小鼠纯合StAR基因敲除等位基因（StAR-/-）的情况下，所有StAR活性都来自StAR转基因，这使我们能够检查它们编码的蛋白质的功能。WT-StAR转基因持续恢复StAR-/-小鼠的生存能力和类固醇生成功能。尽管据报道N47-StAR蛋白在转染的COS细胞和线粒体重建实验中具有活性，但N47-StAR转基因仅在40%的StAR-/-小鼠中恢复了活力。对原发性类固醇生成组织中的脂质沉积的分析揭示了N47-StAR提供的StAR功能等级：在肾上腺皮质和卵巢鞘区可见到丰富的脂质沉积，在间质细胞中可见较轻的沉积。我们的研究结果证实了缺乏线粒体靶向信号的StAR在体内执行一些基本功能的能力，但也证明了与在非甾体源性细胞中获得的体外研究不同的重要功能缺陷。
The steroidogenic acute regulatory protein (StAR) stimulates the regulated production of steroid hormones in the adrenal cortex and gonads by facilitating the delivery of cholesterol to the inner mitochondrial membrane. To explore key aspects of StAR function within bona fide steroidogenic cells, we used a transgenic mouse model to explore the function of StAR proteins in vivo. We first validated this transgenic bacterial artificial chromosome reconstitution system by targeting enhanced green fluorescent protein to steroidogenic cells of the adrenal cortex and gonads. Thereafter, we targeted expression of either wild-type StAR (WT-StAR) or a mutated StAR protein lacking the mitochondrial targeting signal (N47-StAR). In the context of mice homozygous for a StAR knockout allele (StAR-/-), all StAR activity derived from the StAR transgenes, allowing us to examine the function of the proteins that they encode. The WT-StAR transgene consistently restored viability and steroidogenic function to StAR-/- mice. Although the N47-StAR protein was reportedly active in transfected COS cells and mitochondrial reconstitution experiments, the N47-StAR transgene rescued viability in only 40% of StAR-/- mice. Analysis of lipid deposits in the primary steroidogenic tissues revealed a hierarchy of StAR function provided by N47-StAR: florid lipid deposits were seen in the adrenal cortex and ovarian theca region, with milder deposits in the Leydig cells. Our results confirm the ability of StAR lacking its mitochondrial targeting signal to perform some essential functions in vivo but also demonstrate important functional defects that differ from in vitro studies obtained in nonsteroidogenic cells.