Johnston P A, Archer B T, Robinson K, Mignery G A, Jahn R, Südhof T C
Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas 75235.
Neuron. 1991 Jul;7(1):101-9. doi: 10.1016/0896-6273(91)90078-e.
rab3A is a small neuronal GTP-binding protein specifically localized to synaptic vesicles. Membrane-bound rab3A behaves like an intrinsic membrane protein in vitro, but reversibly dissociates from synaptic vesicles after exocytosis in vivo. Here we demonstrate that rab3A is attached to synaptic vesicle membranes by a carboxy-terminal Cys-X-Cys sequence that is posttranslationally modified. This modification is inhibited by compactin in a mevalonate-dependent manner, suggesting that the Cys-X-Cys sequence represents a novel polyisoprenylation sequence. Isolation of a rab3 homolog from D. melanogaster reveals high evolutionary conservation of rab3A, including its carboxy-terminal Cys-X-Cys sequence. The posttranslational modifications of soluble and membrane-bound rab3A are biochemically different, but both require the carboxy-terminal Cys-X-Cys sequence and are faithfully reproduced in nonneuronal cells. Our results suggest that the carboxy-terminal Cys-X-Cys sequence of rab3A is polyisoprenylated and is used as its regulatable membrane anchor. Furthermore, the hydrophobic modification of rab3A and its correct intracellular targeting to synaptic vesicles are independent, presumably consecutive events.
rab3A是一种小的神经元GTP结合蛋白，特异性定位于突触小泡。膜结合的rab3A在体外表现得像一种内在膜蛋白，但在体内胞吐作用后会从突触小泡中可逆地解离。在这里，我们证明rab3A通过一个翻译后修饰的羧基末端Cys-X-Cys序列附着于突触小泡膜。这种修饰被美伐他汀以甲羟戊酸依赖的方式抑制，表明Cys-X-Cys序列代表一种新的多聚异戊二烯化序列。从黑腹果蝇中分离出的rab3同源物显示rab3A具有高度的进化保守性，包括其羧基末端的Cys-X-Cys序列。可溶性和膜结合的rab3A的翻译后修饰在生化性质上有所不同，但两者都需要羧基末端的Cys-X-Cys序列，并且在非神经元细胞中能如实地再现。我们的结果表明，rab3A的羧基末端Cys-X-Cys序列被多聚异戊二烯化，并用作其可调节的膜锚定物。此外，rab3A的疏水修饰及其在细胞内正确靶向突触小泡是独立的，大概是连续的事件。
