From the Department of Molecular Biology and Genetics, Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, New York 14853.
From the Department of Molecular Biology and Genetics, Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, New York 14853
J Biol Chem. 2018 May 4;293(18):6682-6691. doi: 10.1074/jbc.RA117.001318. Epub 2018 Mar 7.
The Golgi complex is the central membrane and protein-sorting station in eukaryotic cells. Activation of Arf (ADP-ribosylation factor) GTPases is essential for vesicle formation via recruitment of cargo adaptors and coat proteins necessary for Golgi trafficking. Arf activation is spatially and temporally regulated by distinct guanine nucleotide exchange factors (GEFs) at different Golgi compartments. The yeast Arf-GEF Sec7 is a conserved and essential activator of Arf1 at the -Golgi network. Sec7 contains a highly conserved regulatory region, the homology upstream of Sec7 (HUS) box, with an unknown mechanistic role. In this study we explore how the HUS box, which is N-terminal to the catalytic domain, acts together with C-terminal regulatory domains in the allosteric activation of Sec7. We report that mutation of the HUS box disrupts positive feedback and allosteric activation of Sec7 by the GTPase Ypt31, a yeast Rab11 homolog. Taken together, our results support a model in which the inter- and intramolecular interactions of the HUS box and the C terminus are necessary for the allosteric activation of Sec7.
高尔基复合体是真核细胞中中央膜和蛋白质分拣站。Arf（ADP-核糖基化因子）GTP 酶的激活对于通过募集货物衔接器和高尔基体运输所需的衣壳蛋白形成囊泡是必不可少的。Arf 的激活通过不同高尔基体隔室中的不同鸟嘌呤核苷酸交换因子（GEF）在空间和时间上受到调节。酵母 Arf-GEF Sec7 是高尔基体网络中 Arf1 的保守且必需的激活剂。Sec7 包含一个高度保守的调节区，上游同源结构域（HUS）盒，其具有未知的机械作用。在这项研究中，我们探讨了 HUS 盒（位于催化结构域的 N 端）如何与 C 端调节结构域一起在 Sec7 的别构激活中发挥作用。我们报告说，HUS 盒的突变破坏了 GTPase Ypt31（酵母 Rab11 同源物）对 Sec7 的正反馈和别构激活。总之，我们的结果支持这样一种模型，即 HUS 盒和 C 端的分子间和分子内相互作用对于 Sec7 的别构激活是必需的。
