Identification and characterization of phosphodiesterases that specifically degrade 3'3'-cyclic GMP-AMP.

Cyclic dinucleotides act as intracellular second messengers, modulating a variety of cellular activities including innate immune activation. Although phosphodiesterases (PDEs) hydrolyzing c-di-GMP and c-di-AMP have been identified, no PDEs for cGAMPs have been reported. Here we identified the first three cGAMP-specific PDEs in V. cholerae (herein designated as V-cGAP1/2/3). V-cGAPs are HD-GYP domain-containing proteins and specifically break 3'3'-cGAMP, but not other forms of cGAMP. 3'3'-cGAMP is first linearized by all three V-cGAPs to produce 5'-pApG, which is further hydrolyzed into 5'-ApG by V-cGAP1. In this two-step reaction, V-cGAP1 functions as both a PDE and a 5'-nucleotidase. In vivo experiments demonstrated that V-cGAPs play non-redundant roles in cGAMP degradation. The high specificity of V-cGAPs on 3'3'-cGAMP suggests the existence of specific PDEs for other cGAMPs, including 2'3'-cGAMP in mammalian cells. The absolute requirement of the GYP motif for 3'3'-cGAMP degradation suggests that HD domain-containing PDEs in eukaryotes are probably unable to hydrolyze cGAMPs. The fact that all V-cGAPs attack 3'3'-cGAMP on one specific phosphodiester bond suggests that PDEs for other cGAMPs would utilize a similar strategy. These results will provide valuable information for identification and characterization of mammalian 2'3'-cGAMP-specific PDEs in future studies.