The transfer of ADP-ribose from NAD(+) to a substrate by ADP-ribosyltransferases, ADP-ribosylation, is a multifunctional posttranslational modification. While many studies have addressed the function of poly-ADP-ribosylation, for example, in DNA repair, signaling, and gene transcription, little is known about the role of mono-ADP-ribosylation. Recent work describing the mono-ADP-ribosyltransferase ARTD10/PARP10 suggests that this enzyme affects apoptosis, NF-κB signaling, and DNA damage repair, at least in part dependent on its activity as mono-ADP-ribosyltransferase. Moreover, the macrodomain-containing proteins MacroD1, MacroD2, and TARG1/C6orf130 were recently described as hydrolases, which remove mono-ADP-ribosylation thus providing evidence that this modification is reversible. In this review, we discuss these novel findings and their broader implications for cell behavior. We suggest functions of ARTD10 in immunity, metabolism, and cancer biology.