Project Summary Mechanistic target of Rapamycin (mTOR) complex 1 (mTORC1) integrates inputs from multiple pathways and senses diverse signals to regulate cell growth, protein translation, and proliferation1,2. Given that spatial compartmentalization can enhance signaling specificity and efficiency3,4, spatial regulation of mTORC1 appears to be critical for this multifaceted signaling complex5 as it has been reported at many subcellular locations6–9. For example, mTORC1 at the lysosome is regulated by both amino acids and growth factors and functions to promote translation and suppress autophagy10. The complex has also been identified at peroxisomes, where mTORC1 responds to reactive oxygen species7. However, the roles of mTORC1 at other subcellular locations where its presence has been reported, in particular in the nucleus, are not well understood. A major limitation in the field is the availability of tools to assess the activity and function of spatially compartmentalized signaling enzymes in living cells. Using a genetically encoded fluorescent biosensor to monitor mTORC1 activity, the Zhang lab has previously discovered a pool of nuclear mTORC1, which has yet to be defined in function. Based on our preliminary data, we hypothesize that nuclear mTORC1 regulates pro-inflammatory transcription. We will test this hypothesis by combining targeted inhibition of mTORC1 with a phosphoproteomics experiment and biochemical assays. Lastly, since genetic mutations activating phosphoinositide 3-kinase(PI3K)/protein kinase B (Akt)/mTOR signaling are prominent alterations in oral squamous cell carcinoma (OSCC)11,12 and mTORC1 inhibition can induce tumor regression13,14, we will study the role of subcellular mTORC1 signaling in the growth and drug resistance of OSCC cells in vitro and in vivo in mice. We hypothesize that inhibiting mTORC1 in the nucleus or the cytosol will differentially effect cell growth and EGFR inhibitor resistance in OSCC. The proposed project will elucidate the function of the previously undefined pool of nuclear mTORC1 and clarify the roles of subcellular mTORC1 in the growth and drug resistance of OSCC.