Modular antibodies reveal DNA damage-induced mono-ADP-ribosylation as a second wave of PARP1 signaling
Edoardo José Longarini, Helen Dauben, Carolina Locatelli, Anne R. Wondisford, Rebecca Smith, Charlotte Muench, Andreas Kolvenbach, Michelle Lee Lynskey, Alexis Pope, Juán José Bonfiglio, Eva Pinto Jurado, Roberta Fajka‐Boja, Thomas Colby, M. Schuller, Ivan Ahel, Gyula Timinszky, Roderick J. O’Sullivan, Sébastien Huet, Ivan Matić
Abstract
PARP1, an established anti-cancer target that regulates many cellular pathways, including DNA repair signaling, has been intensely studied for decades as a poly(ADP-ribosyl)transferase. Although recent studies have revealed the prevalence of mono-ADP-ribosylation upon DNA damage, it was unknown whether this signal plays an active role in the cell or is just a byproduct of poly-ADP-ribosylation. By engineering SpyTag-based modular antibodies for sensitive and flexible detection of mono-ADP-ribosylation, including fluorescence-based sensors for live-cell imaging, we demonstrate that serine mono-ADP-ribosylation constitutes a second wave of PARP1 signaling shaped by the cellular HPF1/PARP1 ratio. Multilevel chromatin proteomics reveals histone mono-ADP-ribosylation readers, including RNF114, a ubiquitin ligase recruited to DNA lesions through a zinc-finger domain, modulating the DNA damage response and telomere maintenance. Our work provides a technological framework for illuminating ADP-ribosylation in a wide range of applications and biological contexts and establishes mono-ADP-ribosylation by HPF1/PARP1 as an important information carrier for cell signaling.