Abstract

Summary 2′3′-cyclic GMP-AMP (2′3′-cGAMP), generated by cyclic GMP-AMP synthase (cGAS) under activation by cytosolic DNA, has a vital role in innate immune response via its receptor protein stimulator of interferon genes (STING) to fight viral infections and tumors. In order to have a complete understanding of biological functions of 2′3′-cGAMP, it is important to find out whether 2′3′-cGAMP has other unrevealed binding proteins present in mammalian cells and executes unknown functions. Here we report the 2′3′-cGAMP-based photoaffinity probes that capture and isolate 2′3′-cGAMP-binding proteins. These probes enable the identification of some potential 2′3′-cGAMP-binding proteins from HeLa cells. EF1A1, an essential protein regulating protein synthesis, is further validated to associate with 2′3′-cGAMP in vitro and in cells to impede protein synthesis. Thus, our studies provide a powerful approach to enable identification of the 2′3′-cGAMP interactome, discover unknown functions of 2′3′-cGAMP, and understand its physiological/pathological roles in tumor immunity and immune-related diseases.