Abstract Mammalian stress-activated protein kinase-interacting protein 1 (mSin1), a core component of the mechanistic target of rapamycin complex 2 (mTORC2), is less well characterized than other mTOR subunits. In particular, the mechanisms regulating its phosphorylation and its functional contribution to mTOR signaling remain poorly defined. Conflicting reports suggest that phosphorylation of mSin1 at T86 is mediated either by Akt or by ribosomal protein S6 kinase 1 (S6K1), highlighting uncertainty regarding its upstream regulation. These discrepancies underscore the need for deeper investigation into the kinases and pathways governing mSin1 phosphorylation. We found that rapamycin treatment decreases overall phosphorylation while paradoxically increasing T86 phosphorylation of mSin1 across multiple normal and cancer cell lines. These effects are independent of mTORC1/mTORC2 and downstream effectors (S6K1 and Akt) yet requires mTOR kinase activity and the subunit mLST8 (mammalian lethal with SEC13 protein 8). Since mTOR acts as a nutrient sensor, we also examined the effect of nutrient availability on mSin1 phosphorylation. Prolonged glucose deprivation, but not amino acid deprivation, markedly decreased mSin1 phosphorylation in HeLa cells, which was restored upon glucose repletion. These effects were not observed in Rh30, VSMC, and MEF cells. Collectively, our findings suggest that that rapamycin inhibits mSin1 phosphorylation by targeting a novel mTOR complex and might be associated with glucose metabolism in some contexts. Further research is needed to define the exact phosphorylation sites and their significance as well as the new mTOR complex regulating these phosphorylation sites. Citation Format: Samira Mahmoudi, Bing Cheng, Yan Luo, Lei Liu, Shile Huang, . The mystery behind mSin1 phosphorylation abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 3316.
Mahmoudi et al. (2026) studied this question.