Family-wide analysis of human macrodomains reveals novel activities and identifies PARG as most efficient ADPr-RNA hydrolase

ADP-ribosylation is well-known as protein posttranslational modification and was recently also identified as RNA posttranscriptional modification. When macrodomain proteins were identified as protein ADP-ribosylhydrolases, several ADP-ribosylation substrates were not yet identified. Therefore, the majority of macrodomain-containing proteins have not been tested towards these additional substrates and were considered to be inactive. Here, we compare in vitro activities of the human macrodomains on a range of ADP-ribosylated substrates. We confirm recent findings that PARP9macro1 and PARP14macro1 can remove ADP-ribose from acidic residues and provide evidence that also PARP14macro2 and PARP15macro2 can function as ADP-ribosylhydrolases. In addition, we find that both PARP9macro1 and PARP14macro1 are active as ADPr-RNA decapping protein domains. Notwithstanding these in vitro activities, our data furthermore indicate that in HEK293 cells, PARG is the major ADPr-RNA decapping enzyme. Our findings thus expand the spectrum of known catalytic activities of human macrodomains and demonstrate their different efficiencies towards nucleic acid substrates.