Abstract
Poly-ADP-ribose polymerase (PARP)-family ADP-ribosyltransferases function in various signaling pathways, predominantly in the nucleus and cytosol. Although PARP inhibitors are in clinical practice for cancer therapy, the enzymatic activities of individual PARP family members are yet insufficiently understood. We studied PARP10, a mono-ADP-ribosyltransferase and potential drug target. Using acid-urea gel electrophoresis, we found that the isolated catalytic domain of PARP10 auto-ADP-ribosylates (MARylates) at eight or more acceptor residues. We isolated individual species with either singular or several modifications and then analyzed them by mass spectrometry. The results confirmed multi-site MARylation in a random order and identified four acceptor residues. The mutagenesis of singular acceptor residues had a minor impact on the overall auto-MARylation level and no effect on the MARylation of histone H3.1. Together, our results suggest that PARP10 automodification may have functions in the regulation of intramolecular or partner binding events, rather than of its enzymatic catalysis. This contributes to a better understanding of PARP10 functions, and, in the long run, to gauging the consequences of PARP inhibitor actions.
Highlights
IntroductionWhile the human genome contains an estimated 25,000 genes, the human proteome consists of more than one million different polypeptides [1]
Single genes encode multiple differentially functional proteins
When PARP10 auto-MARylation reaction mixtures were separated by acid-urea gel electrophoresis (Appendix A), unmodified and modified proteins were resolved (Figure 1)
Summary
While the human genome contains an estimated 25,000 genes, the human proteome consists of more than one million different polypeptides [1] This increase in complexity is possible thanks to, among other mechanisms, protein posttranslational modifications (PTMs), covalent alterations that allow for the swift modulation of enzymatic activities and subcellular localizations. These enzymes catalyze the transfer of a single unit of ADP-ribose to their target, consuming a molecule of nicotinamide adenine dinucleotide (NAD+ ) in the process. They can transfer ADP-ribose to a wide array of targets, including glutamate, aspartate, serine, lysine, and arginine side chains in peptides and proteins [5,6], and to phosphorylated ends of RNA [7]. The arguably most well-studied enzymes in this sub-family are PARP10/ARTD10 and PARP14/ARTD8/BAL2; but we are only beginning to understand their substrate specificities and biological implementations
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
