Department of Molecular Biology and Ecology of Plants, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv 69978, Israel.
Curr Biol. 2010 May 25;20(10):914-20. doi: 10.1016/j.cub.2010.03.057. Epub 2010 May 6.
Rho GTPases are master regulators of cell polarity. For their function, Rhos must associate with discrete plasma membrane domains. Rho of Plants (ROPs) or RACs comprise a single family. Prenylation and S-acylation of hypervariable domain cysteines of Ras and Rho GTPases are required for their function; however, lipid modifications in the G domain have never been reported. Reversible S-acylation involves the attachment of palmitate (C16:0) or other saturated lipids to cysteines through a thioester linkage and was implicated in the regulation of signaling. Here we show that transient S-acylation of Arabidopsis AtROP6 takes place on two conserved G domain cysteine residues, C21 and C156. C21 is relatively exposed and is accessible for modification, but C156 is not, implying that its S-acylation involves a conformational change. Fluorescence recovery after photobleaching beam-size analysis shows that S-acylation of AtROP6 regulates its membrane-association dynamics, and detergent-solubilization studies indicate that it regulates AtROP6 association with lipid rafts. Site-specific acylation-deficient AtROP6 mutants can bind and hydrolyze GTP but display compromised effects on polar cell growth, endocytic uptake of the tracer dye FM4-64, and distribution of reactive oxygen species. These data reveal an S-acylation switch that regulates Rho signaling.
Rho GTPases 是细胞极性的主要调节因子。为了发挥其功能，Rhos 必须与离散的质膜域结合。植物 Rho（ROPs）或 Rac 构成了一个单一的家族。Ras 和 Rho GTPases 超变域半胱氨酸的prenylation 和 S-acylation 对于其功能是必需的；然而，G 结构域中的脂质修饰从未被报道过。可逆的 S-acylation 涉及通过硫酯键将棕榈酸（C16:0）或其他饱和脂质附着到半胱氨酸上，并且与信号转导的调节有关。在这里，我们表明拟南芥 AtROP6 的瞬时 S-acylation 发生在两个保守的 G 结构域半胱氨酸残基 C21 和 C156 上。C21 相对暴露且易于修饰，但 C156 则不然，这表明其 S-acylation 涉及构象变化。光漂白后荧光恢复光束尺寸分析表明，AtROP6 的 S-acylation 调节其膜结合动力学，并且去污剂溶解研究表明它调节 AtROP6 与脂筏的结合。具有特定位置的去酰化缺陷 AtROP6 突变体可以结合并水解 GTP，但对极性细胞生长、示踪染料 FM4-64 的内吞摄取以及活性氧的分布的影响受损。这些数据揭示了调节 Rho 信号的 S-acylation 开关。
