DEPTOR regulates nucleus pulposus cell senescence through the mTORC1/S6K1/ATG1 pathway to alleviate intervertebral disk degeneration

Abstract This study aimed to determine the molecular mechanisms by which the DEP domain-containing mTOR-interacting protein (DEPTOR) regulates the senescence of nucleus pulposus (NP) cells (NPCs), alleviating intervertebral disk degeneration (IDD). This study investigated how DEPTOR regulates the mechanistic target of rapamycin complex 1 (mTORC1)/S6 kinase beta-1 (S6K1)/autophagy-related gene 1 (ATG1) pathway to regulate senescence-associated secretory phenotype (SASP) and cellular autophagy in NPCs. Isobaric tags for relative and absolute quantitation was used to measure the differences in protein expression between degenerated and normal intervertebral disk tissues. Western blotting and immunofluorescence were used to quantify DEPTOR levels in NP tissues. DEPTOR was overexpressed in vitro, and changes in autophagy and SASP were monitored to determine its effects on NPCs. Moreover, lentiviral overexpression of S6K1 (LV-S6K1) and siRNA-mediated knockdown of ATG1 (ATG1-ShRNA) in both in vitro and in vivo models were used to verify whether DEPTOR stimulates autophagy in NPCs via ATG1 and inhibits SASP through S6K1. The results demonstrated that degenerated intervertebral disks had lower DEPTOR levels. Matrix metalloproteinases, inflammatory cytokines, chemokines, and aging-related proteins were downregulated when DEPTOR was overexpressed in NPCs. Furthermore, autophagic activity was stimulated, SASP secretion was inhibited, and extracellular matrix synthesis was increased. ATG1 knockdown decreased the capacity of DEPTOR to promote cellular autophagy and inhibit SASP, whereas S6K1 overexpression diminished DEPTOR-mediated SASP inhibition. DEPTOR attenuates IDD by inhibiting SASP secretion via the mTORC1/S6K1 pathway and promoting autophagy in NPCs via the mTORC1/ATG1 pathway.