CRISPR screening identifies GNPTAB as a noncanonical STING activator driving cellular senescence.

Cellular senescence is accompanied by profound lysosomal alterations, yet whether lysosome-associated factors actively drive aging remains unclear. Through a focused CRISPR/Cas9 screen in human mesenchymal progenitor cells (hMPCs), we identified N-acetylglucosamine-1-phosphotransferase subunits alpha and beta (GNPTAB), an enzyme responsible for lysosomal hydrolase targeting, as a potent regulator of cellular senescence. Genetic ablation of GNPTAB attenuated senescence, whereas its overexpression accelerated senescence. This pro-senescent function occurred independently of GNPTAB's canonical enzymatic role. Instead, GNPTAB binds to the innate immune adaptor stimulator of interferon genes (STING) via a specific interface (E1119), leading to activation of STING and its downstream TANK-binding kinase 1 (TBK1), as well as pro-inflammatory gene expression. A STING-binding-deficient GNPTAB mutant (E1119A) preserved canonical lysosomal functions but failed to induce senescence, while STING depletion abolished GNPTAB-driven senescence. Together, these findings uncover a new signaling pathway wherein GNPTAB engages STING to facilitate its activation, nominating this interface as a potential target for mitigating age-related cellular dysfunction.