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The Loss of Lam2 and Npr2-Npr3 Diminishes the Vacuolar Localization of Gtr1-Gtr2 and Disinhibits TORC1 Activity in Fission Yeast.

PLoS ONE. 2016 May 26;11(5):e0156239. eCollection 2016
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摘要


In mammalian cells, mTORC1 activity is regulated by Rag GTPases. It is thought that the Ragulator complex and the GATOR (GAP activity towards Rags) complex regulate RagA/B as its GDP/GTP exchange factor (GEF) and GTPase-activating protein (GAP), respectively. However, the functions of components in these complexes remain elusive. Using fission yeast as a model organism, here we found that the loss of Lam2 (SPBC1778.05c), a homolog of a Ragulator component LAMTOR2, as well as the loss of Gtr1 or Gtr2 phenocopies the loss of Npr2 or Npr3, homologs of GATOR components Nprl2 or Nprl3, respectively. These phenotypes were rescued by TORC1 inhibition using pharmacological or genetic means, and the loss of Lam2, Gtr1, Gtr2, Npr2 or Npr3 disinhibited TORC1 activity under nitrogen depletion, as measured by Rps6 phosphorylation. Consistently, overexpression of GDP-locked Gtr1S20L or GTP-locked Gtr2Q60L, which suppress TORC1 activity in budding yeast, rescued the growth defect of Δgtr1 cells or Δgtr2 cells, respectively, and the loss of Lam2, Npr2 or Npr3 similarly diminished the vacuolar localization and the protein levels of Gtr1 and Gtr2. Furthermore, Lam2 physically interacted with Npr2 and Gtr1. These findings suggest that Lam2 and Npr2-Npr3 function together as a tether for GDP-bound Gtr1 to the vacuolar membrane, thereby suppressing TORC1 activity for multiple cellular functions.

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