Poster Presentation 51st Lorne Proteins Conference 2026

Structure of the lysosomal KICSTOR-GATOR1-SAMTOR nutrient-sensing supercomplex (#422)

Chris J Lupton 1 , Charles Bayly-Jones 1 2 , Shuqi Dong 3 , Terrance Lam 3 , Wentong Luo 1 , Gareth D Jones 1 , Chantel Mastos 3 , Nicholas J Frescher 1 , San Sui Lim 1 , Alastair C Keen 3 , Luke E Formosa 1 , Hariprasad Venugopal 4 , Tyler Chang 1 , Michelle L Halls 3 , Andrew M Ellisdon 1
  1. Biomedical Discovery Institute, Monash University, Melbourne, Victoria, Australia
  2. School of Chemistry, University of Sydney, Sydney, New South Wales, Australia
  3. Drug Discovery Biology, Monash Institute of Pharmaceutical Sciences, Melbourne, Victoria, Australia
  4. Ramaciotti Centre for Cryo-Electron Microscopy, Monash University, Melbourne, Victoria, Australia

The guanosine triphosphate (GTP)-bound state of the heterodimeric Rag GTPases functions as a molecular switch regulating mechanistic target of rapamycin complex 1 (mTORC1) activation at the lysosome downstream of amino acid fluctuations. Under low amino acid conditions, GTPase-activating protein (GAP) activity toward Rags 1 (GATOR1) promotes RagA GTP hydrolysis, preventing mTORC1 activation. KICSTOR recruits and regulates GATOR1 at the lysosome by undefined mechanisms. Here, we resolve the KICSTOR-GATOR1 structure, revealing a striking ∼60-nm crescent-shaped assembly. GATOR1 anchors to KICSTOR via an extensive interface, and mutations that disrupt this interaction impair mTORC1 regulation. The S-adenosylmethionine sensor SAMTOR binds KICSTOR in a manner incompatible with metabolite binding, providing structural insight into methionine sensing via SAMTOR-KICSTOR association. We discover that KICSTOR and GATOR1 form a dimeric supercomplex. This assembly restricts GATOR1 to an orientation that favors the low-affinity active GAP mode of Rag GTPase engagement while sterically restricting access to the high-affinity inhibitory mode, consistent with a model of an active lysosomal GATOR1 docking complex.