Chemical Proteomics of the Tumor Suppressor Fhit Covalently Bound to the Cofactor Ap₃A Elucidates Its Inhibitory Action on Translation

The tumor suppressor protein fragile histidine triad (Fhit) is known to be associated with genomic instability and apoptosis. The tumor-suppressive function of Fhit depends on the interaction with the alarmone diadenosine triphosphate (Ap₃A), a noncanonical nucleotide whose concentration increases upon cellular stress. How the Fhit–Ap₃A complex exerts its signaling function is unknown. Here, guided by a chemical proteomics approach employing a synthetic stable Fhit–Ap₃A complex, we found that the Fhit–Ap₃A complex, but not Fhit or Ap₃A alone, impedes translation. Our findings provide a mechanistic model in which Fhit translocates from the nucleolus into the cytosol upon stress to form an Fhit–Ap₃A complex. The Fhit–Ap₃A complex impedes translation both in vitro and in vivo, resulting in reduced cell viability. Overall, our findings provide a mechanistic model by which the tumor suppressor Fhit collaborates with the alarmone Ap₃A to regulate cellular proliferation.