Redundant electrostatic interactions between GATOR1 and the Rag GTPase heterodimer drive efficient amino acid sensing in human cells.
Redundant electrostatic interactions between GATOR1 and the Rag GTPase heterodimer drive efficient amino acid sensing in human cells.
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DOI：
10.1016/j.jbc.2023.104880
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发表时间：
2023-07
影响因子：
4.8
通讯作者：
Shen, Kuang
中科院分区：
文献类型：
--
作者：
Doxsey, Dylan D.;Tettoni, Steven D.;Egri, Shawn B.;Shen, Kuang
细胞需要协调营养供应与其生长和增殖。在真核细胞中，这种协调是由雷帕霉素复合物1（mTORC 1）途径的机制靶点介导的。mTORC 1的激活受两个GT3单位调节，Rag GT3异二聚体和Rheb GT3。RagA-RagC异源二聚体控制mTORC 1的亚细胞定位，其核苷酸加载状态受到上游调控因子包括氨基酸传感器的严格控制。Rag GT3异二聚体的关键负调节因子是GATOR 1。在缺乏氨基酸的情况下，GATOR 1通过RagA亚基刺激GTP水解以关闭mTORC 1信号传导。尽管GATOR 1对RagA具有酶特异性，但最近的人GATOR 1-Rag-Ragulator复合物的cryo-EM结构模型揭示了GATOR 1的亚基Depdc 5与RagC之间的意想不到的界面。目前，还没有这个接口的功能特性，我们也不知道它的生物学相关性。在这里，结合结构-功能分析，酶动力学测量和基于细胞的信号传导测定，我们确定了Depdc 5和RagC之间的关键静电相互作用。这种相互作用由Depdc 5上带正电荷的Arg-1407残基和RagC侧面上带负电荷的残基的补丁介导。消除这种相互作用会损害GATOR 1差距活性和细胞对氨基酸去除的反应。我们的研究结果揭示了GATOR 1如何协调Rag GT3异源二聚体的核苷酸加载状态，从而精确地控制在没有氨基酸的情况下的细胞行为。
Cells need to coordinate nutrient availability with their growth and proliferation. In eukaryotic cells, this coordination is mediated by the mechanistic target of the rapamycin complex 1 (mTORC1) pathway. mTORC1 activation is regulated by two GTPase units, the Rag GTPase heterodimer and the Rheb GTPase. The RagA-RagC heterodimer controls the subcellular localization of mTORC1, and its nucleotide loading states are strictly controlled by upstream regulators including amino acid sensors. A critical negative regulator of the Rag GTPase heterodimer is GATOR1. In the absence of amino acids, GATOR1 stimulates GTP hydrolysis by the RagA subunit to turn off mTORC1 signaling. Despite the enzymatic specificity of GATOR1 to RagA, a recent cryo-EM structural model of the human GATOR1-Rag-Ragulator complex reveals an unexpected interface between Depdc5, a subunit of GATOR1, and RagC. Currently, there is no functional characterization of this interface, nor do we know its biological relevance. Here, combining structure-function analysis, enzymatic kinetic measurements, and cell-based signaling assays, we identified a critical electrostatic interaction between Depdc5 and RagC. This interaction is mediated by the positively charged Arg-1407 residue on Depdc5 and a patch of negatively charged residues on the lateral side of RagC. Abrogating this interaction impairs the GAP activity of GATOR1 and cellular response to amino acid withdrawal. Our results reveal how GATOR1 coordinates the nucleotide loading states of the Rag GTPase heterodimer, and thus precisely controls cellular behavior in the absence of amino acids.
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影响因子：
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作者：
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