Divergent signaling profiles in mTOR gain-of-function Smith-Kingsmore syndrome (SKS) and TSC2 deficiency.

Smith-Kingsmore syndrome (SKS) is a rare neurodevelopmental disorder caused by gain-of-function mutations in MTOR, yet whether these mutations phenocopy TSC2 loss or establish a distinct signaling state remains unclear. Using quantitative proteomics, phosphoproteomics, and transcriptomics in isogenic cell models of SKS (MTOR Δ4aa ), TSC2 loss (TSC2 -/-), and wild-type controls under glucose depletion and refeeding, we find that MTOR Δ4aa and TSC2 -/- cells occupy fundamentally distinct regulatory states. TSC2 -/- cells exhibit broad anabolic remodeling and a transcriptional program dominated by NF-κB- and STAT-driven inflammatory responses. MTOR Δ4aa cells instead display enrichment of nuclear and RNA processing programs, E2F/MYC-driven transcription, and a constrained proteomic dynamic range across nutrient states. Phosphoproteomic analysis of MTOR Δ4aa reveals rerouting of nutrient-responsive signaling toward MAPK/ERK- and Ca2+/CaMK-dependent pathways with limited canonical mTORC1/S6K1 engagement. These findings establish SKS as a signaling rewiring disorder distinct from classical mTORC1 hyperactivation, with implications for therapeutic targeting.