Electrical properties of plasma membrane modulate subcellular distribution of K-Ras
Electrical properties of plasma membrane modulate subcellular distribution of K-Ras
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DOI：
10.1111/j.1742-4658.2007.05758.x
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发表时间：
2007-05-01
期刊：
影响因子：
5.4
通讯作者：
Daniotti, Jose L.
中科院分区：
文献类型：
--
作者：
Gomez, Guillermo A.;Daniotti, Jose L.
K-Ras是一种小G蛋白，主要位于质膜的内小叶。该蛋白的膜靶向信号由六个连续赖氨酸和法尼基化半胱氨酸的多元C-末端序列组成。模型系统中的生物物理研究结果表明，疏水和静电相互作用是负责K-Ras的膜结合特性。为了在细胞系统中验证这一假设，我们首先在体外评估了电解质对K-Ras膜结合特性的影响。结果表明K-Ras与膜中阴离子脂质结合的电性和可逆性。接下来，我们研究了膜结合和K-Ras的亚细胞分布后，破坏的质膜和离子梯度的外部和内部小叶的电性能。从外部质膜的唾液酸的去除引起的K-Ras的再分配再循环内体。通过消耗细胞ATP抑制质膜上的聚磷酸肌醇合成，导致K-Ras在类似的亚细胞内重新分布。用改变跨膜电位的离子载体处理细胞引起K-Ras向细胞质和内膜的重新分布。Ca ~（2+）离子载体与K ~+离子载体相比，引起K-Ras向内膜的更广泛的再分布。总之，这些结果揭示了K-Ras和细胞膜之间相互作用的动态性质，并表明K-Ras的亚细胞分布是由蛋白质的多碱基区域与带负电荷的膜的静电相互作用驱动的。
K-Ras is a small G-protein, localized mainly at the inner leaflet of the plasma membrane. The membrane targeting signal of this protein consists of a polybasic C-terminal sequence of six contiguous lysines and a farnesylated cysteine. Results from biophysical studies in model systems suggest that hydrophobic and electrostatic interactions are responsible for the membrane binding properties of K-Ras. To test this hypothesis in a cellular system, we first evaluated in vitro the effect of electrolytes on K-Ras membrane binding properties. Results demonstrated the electrical and reversible nature of K-Ras binding to anionic lipids in membranes. We next investigated membrane binding and subcellular distribution of K-Ras after disruption of the electrical properties of the outer and inner leaflets of plasma membrane and ionic gradients through it. Removal of sialic acid from the outer plasma membrane caused a redistribution of K-Ras to recycling endosomes. Inhibition of polyphosphoinositide synthesis at the plasma membrane, by depletion of cellular ATP, resulted in a similar subcellular redistribution of K-Ras. Treatment of cells with ionophores that modify transmembrane potential caused a redistribution of K-Ras to cytoplasm and endomembranes. Ca2+ ionophores, compared to K+ ionophores, caused a much broader redistribution of K-Ras to endomembranes. Taken together, these results reveal the dynamic nature of interactions between K-Ras and cellular membranes, and indicate that subcellular distribution of K-Ras is driven by electrostatic interaction of the polybasic region of the protein with negatively charged membranes.