Hierarchical inhibition of mTORC1 by glucose starvation-triggered AXIN lysosomal translocation and by AMPK

Abstract When glucose is replete, mTORC1 is active and anchored to the lysosomal surface via the two GTPases, RAG and Rheb, which are regulated by Ragulator and TSC2, respectively. When glucose is low, aldolase senses low fructose-1,6-bisphosphate and promotes the translocation of AXIN−LKB1 to the lysosomal surface, which leads to the activation of AMPK and the inhibition of RAGs, sundering mTORC1 from the lysosome and causing its inactivation. AMPK can also inactivate mTORC1 by phosphorylating Raptor and TSC2. However, the hierarchy of AXIN- and AMPK-mediated inhibition of mTORC1 remains poorly defined. Here, we show that AXIN translocation does not require AMPK expression or activity. In glucose starvation conditions, knockout of AXIN extended the half-life of mTORC1 inhibition from 15 min to 60 min, whereas knockout of AMPK only extended it to 30 min. RAGB GTP (constitutively active RAGB) almost entirely blocked the lysosomal dissociation and inhibition of mTORC1 under glucose starvation, but it did not inhibit AMPK, indicating that under these conditions it is AXIN lysosomal translocation that inhibits mTORC1, and it does so via inhibition of RAGs. AICAR, a mimetic of AMP, which activates both cytosolic AMPK and lysosomal AMPK, fully inhibited mTORC1 even when it is stably anchored to the lysosome by RAGB GTP, whereas glucose starvation mildly inhibited such anchored mTORC1. Together, we demonstrate that the lysosomal translocation of AXIN plays a primary role in glucose starvation-triggered inhibition of mTORC1 by inhibiting RAGs, and that AMPK activity inhibits mTORC1 through phosphorylating Raptor and TSC2, especially under severe stress.