GATOR1(GAP活性TOwardRag1)是一种进化保守的GTP酶激活蛋白复合物,可控制mTORC1(哺乳动物雷帕霉素靶标复合物1)对细胞中氨基酸可用性的反应。GATOR1亚基的基因突变,NPRL2(氮通透酶调节因子样2),NPRL3(氮通透酶调节因子样3),和DEPDC5(含有5个DEP域),与人类癫痫有关;然而,这些突变对GATOR1功能和mTORC1调控的具体影响尚不清楚.在这里,我们报道了GATOR1的NPRL2亚基,NPRL2-L105P,-T110S,和-D214H,增加细胞中基础mTORC1信号转导。值得注意的是,我们发现NPRL2-L105P是一种功能缺失突变,它破坏了蛋白质与NPRL3和DEPDC5的相互作用,损害了GATOR1复合物的组装,甚至在氨基酸剥夺条件下也能产生高mTORC1活性.此外,我们的研究表明,GATOR1复合物对于mTORC1的快速和强大抑制是必需的,以响应生长因子的戒断或磷脂酰肌醇-3激酶(PI3K)的药理抑制。在没有GATOR1复合体的情况下,细胞难以抑制PI3K依赖性的mTORC1抑制,允许持续翻译和限制TFEB的核定位,由mTORC1调节的转录因子。总的来说,我们的结果表明,NPRL2中的癫痫相关突变可以阻断GATOR1复合物的组装,并在存在或不存在氨基酸的情况下,限制典型PI3K依赖性生长因子信号对mTORC1的适当调节.
GATOR1 (GAP Activity TOward Rag 1) is an evolutionarily conserved GTPase-activating protein complex that controls the activity of mTORC1 (mammalian Target Of Rapamycin Complex 1) in response to amino acid availability in cells. Genetic mutations in the GATOR1 subunits, NPRL2 (nitrogen permease regulator-like 2), NPRL3 (nitrogen permease regulator-like 3), and DEPDC5 (DEP domain containing 5), have been associated with epilepsy in humans; however, the specific effects of these mutations on GATOR1 function and mTORC1 regulation are not well understood. Herein, we report that epilepsy-linked mutations in the NPRL2 subunit of GATOR1, NPRL2-L105P, -T110S, and -D214H, increase basal mTORC1 signal transduction in cells. Notably, we show that NPRL2-L105P is a loss-of-function mutation that disrupts protein interactions with NPRL3 and DEPDC5, impairing GATOR1 complex assembly and resulting in high mTORC1 activity even under conditions of amino acid deprivation. Furthermore, our studies reveal that the GATOR1 complex is necessary for the rapid and robust inhibition of mTORC1 in response to growth factor withdrawal or pharmacological inhibition of phosphatidylinositol-3 kinase (PI3K). In the absence of the GATOR1 complex, cells are refractory to PI3K-dependent inhibition of mTORC1, permitting sustained translation and restricting the nuclear localization of TFEB, a transcription factor regulated by mTORC1. Collectively, our results show that epilepsy-linked mutations in NPRL2 can block GATOR1 complex assembly and restrict the appropriate regulation of mTORC1 by canonical PI3K-dependent growth factor signaling in the presence or absence of amino acids.