Regulation of the Rheb GTPase via GATOR1 Complex.

Area of Science:

• Cellular Biology
• Molecular Signaling
• Metabolism Regulation

Background:

• The mechanistic target of rapamycin complex 1 (mTORC1) pathway integrates amino acid and growth factor signals to control cell growth and metabolism.
• Two key upstream regulators are the Rag GTPases (amino acid sensing) and Rheb GTPase (growth factor sensing), inhibited by GATOR1 and TSC complexes, respectively.
• The precise upstream crosstalk and coordination between these branches, especially in organisms lacking the TSC complex, remain poorly understood.

Purpose of the Study:

• To investigate potential upstream crosstalk between the Rag and Rheb GTPase branches of mTORC1 signaling.
• To identify molecular mechanisms coordinating mTORC1 regulation in organisms lacking the TSC complex, such as C. elegans.
• To elucidate the direct physical interactions governing the integration of nutrient and growth factor inputs into mTORC1 signaling.

Main Methods:

• Unbiased quantitative proteomics in C. elegans to identify novel interactors of the Rheb ortholog, RHEB-1.
• Biochemical validation in human cells, including pulldown assays and analysis of Rheb mutants with altered nucleotide-binding states.
• Structural modeling using AlphaFold3 to predict the binding interface and mode of interaction between Rheb and GATOR1 subunits.

Main Results:

• The GATOR1 complex was identified as a novel interactor of RHEB-1 in C. elegans.
• Nucleotide-free Rheb directly binds to the Nprl2-Nprl3 subunits of the GATOR1 complex in human cells, an interaction induced by nutrient stress.
• Structural modeling suggests Rheb binds GATOR1 via a non-catalytic mechanism at a distinct site from the Rag GTPase GAP activity.

Conclusions:

• A conserved, nutrient-regulated physical interaction exists between Rheb and the GATOR1 complex (Nprl2/3 subunits).
• This interaction represents a previously unrecognized point of convergence between the growth factor and amino acid sensing branches of mTORC1 signaling.
• The findings provide a direct molecular link between the Rag and Rheb pathways, explaining how nutrient stress fine-tunes mTORC1 activity.