Gtr/Ego-independent TORC1 activation is achieved through a glutamine-sensitive interaction with Pib2 on the vacuolar membrane

TORC1 is a central regulator of cell growth in response to amino acids. The role of the evolutionarily conserved Gtr/Rag pathway in the regulation of TORC1 is well-established. Recent genetic studies suggest that an additional regulatory pathway, depending on the activity of Pib2, plays a role in TORC1 activation independently of the Gtr/Rag pathway. However, the interplay between the Pib2 pathway and the Gtr/Rag pathway remains unclear. In this study, we show that Pib2 and Gtr/Ego form distinct complexes with TORC1 in a mutually exclusive manner, implying dedicated functional relationships between TORC1 and Pib2 or Gtr/Rag in response to specific amino acids. Furthermore, simultaneous depletion of Pib2 and the Gtr/Ego system abolishes TORC1 activity and completely compromises the vacuolar localization of TORC1. Thus, the amino acid-dependent activation of TORC1 is achieved through the Pib2 and Gtr/Ego pathways alone. Finally, we show that glutamine induces a dose-dependent increase in Pib2-TORC1 complex formation, and that glutamine binds directly to the Pib2 complex. These data provide strong preliminary evidence for Pib2 functioning as a putative glutamine sensor in the regulation of TORC1.

雷帕霉素靶蛋白复合物1（TORC1）是响应氨基酸信号调控细胞生长的核心调控因子。进化上保守的Gtr/Rag通路（Gtr/Rag pathway）对TORC1的调控作用已得到充分证实。近期的遗传学研究表明，另一条依赖于Pib2活性的调控通路可独立于Gtr/Rag通路参与TORC1的激活过程。然而，Pib2通路与Gtr/Rag通路之间的相互作用机制仍不明确。本研究发现，Pib2与Gtr/Ego分别以互斥的方式与TORC1形成不同的复合物，这暗示在响应特定氨基酸信号时，TORC1与Pib2或Gtr/Rag通路之间存在专属的功能关联。此外，同时敲除Pib2与Gtr/Ego系统会完全消除TORC1的活性，并彻底破坏TORC1的液泡定位。由此可见，氨基酸依赖的TORC1激活仅通过Pib2与Gtr/Ego两条通路即可实现。最后，我们发现谷氨酰胺可呈剂量依赖性地促进Pib2-TORC1复合物的形成，且谷氨酰胺可直接结合至Pib2复合物中。上述数据为Pib2作为调控TORC1的潜在谷氨酰胺感受器提供了强有力的初步实验证据。