Suppression of ITPK1 and IPMK activities impairs mTORC1 signaling due to accelerated signal termination and prevents pancreatic β-cell metabolic adaptation to excess glucose

Abstract

mTORC1 integrates growth factor and nutrient signals to regulate cellular metabolism, yet there are no metabolites known to directly regulate mTORC1 activity in cells. Cryo-EM studies revealed that inositol hexakisphosphate (IP6) associates with the FAT domain of mTOR, suggesting that inositol phosphates may directly modulate mTOR activity. We previously showed that higher-order inositol phosphates enhance mTORC1 kinase activity and stability in vitro. Here, we investigated whether inositol phosphate metabolism regulates mTORC1 signaling in HEK293 and pancreatic {beta}-cells. Suppression or acute inhibition of inositol phosphate multikinase (IPMK) selectively reduced cellular IP5 levels without altering IP6 and resulted in reduced basal and insulin-stimulated mTORC1 signaling, without decreasing mTORC1 complex stability or lysosomal localization. ITPK1 knockdown phenocopied IPMK inhibition by reducing IP5 and basal mTORC1. Combined inhibition of IPMK and ITPK1 nearly abolished IP5 and reduced IP6, demonstrating that these enzymes compensate to supply IP5 for IP6 synthesis. Importantly, depletion of IP5 did not impair PI3K/Akt activation but accelerated termination of the mTORC1 signal, indicating a role for IP5 in stabilizing the active mTORC1 complex. Reduction of IP5 levels did not prevent insulin- or glucose-induced mTORC1 activation, revealing that inositol phosphates primarily regulate signal persistence rather than initiation. Lastly, we show that suppression of IPMK or ITPK1 in {beta}-cells delayed the mTORC1-dependent metabolic acceleration in response to excess glucose. Together, these findings identify higher order inositol phosphates as metabolic regulators that prolong mTORC1 activity, providing a potential strategy for modulating mTORC1 signaling in cells.