SummaryALC1, a novel PARP1-stimulated chromatin-remodelling enzyme promotes DNA repair.Post-translational modifications play key roles in orchestrating chromatin plasticity. Although various chromatin-remodelling enzymes have been described that respond to specific histone modifications, little is known about the role of poly(ADP-ribose) in chromatin remodelling. Here, we identify a novel chromatin-remodelling enzyme, ALC1 (Amplified in Liver Cancer 1), that is specifically regulated by poly(ADP-ribosyl) ation. ALC1 binds poly(ADP-ribose) via a C-terminal Macro domain and catalyzes PARP1-stimulated nucleosome sliding, conferred by an N-terminal ISWI-related helicase core. Our results define ALC1 as a novel DNA damage-response protein, whose role in this process is sustained by its association with known DNA repair factors and its rapid poly(ADP-ribose)-dependent recruitment to DNA damage sites. Furthermore, we show that depletion or overexpression of ALC1 results in sensitivity to DNA-damaging agents. Collectively, these results provide new insights into the mechanisms by which poly(ADP-ribose) regulates DNA repair.The restricted accessibility of DNA within chromatin presents a barrier to DNA manipulations that require direct protein-DNA interactions (1-3). Processes such as transcription, repair and replication that require efficient DNA recognition are therefore dependent on the appropriate modulation of chromatin structure. Chromatin relaxation is a critical event that occurs during DNA repair and is associated with post-translational poly(ADP-ribose) (PAR) modification (4). PAR is synthesized in a reaction that utilizes NAD+ as a substrate by the PARP family of enzymes, of which PARP1 (and to a lesser extent PARP2) respond to DNA strand breaks (5-7). As a consequence of poly(ADP-

The poly(ADP-ribose) polymerase (PARP) protein family generates ADP-ribose (ADPr) modifications onto target proteins using NAD+ as substrate. Based on the composition of three NAD+ coordinating amino acids, the H-Y-E motif, each PARP is predicted to generate either poly(ADP-ribose) (PAR) or mono(ADP-ribose) (MAR). However, the reaction product of each PARP has not been clearly defined, and is an important priority since PAR and MAR function via distinct mechanisms. Here we show that the majority of PARPs generate MAR, not PAR, and demonstrate that the H-Y-E motif is not the sole indicator of PARP activity. We identify automodification sites on seven PARPs, and demonstrate that MAR and PAR generating PARPs modify similar amino acids, suggesting that the sequence and structural constraints limiting PARPs to MAR synthesis do not limit their ability to modify canonical amino acid targets. In addition, we identify cysteine as a novel amino acid target for ADP-ribosylation on PARPs.

Posttranslational modification of proteins by poly(ADP-ribosyl)ation regulates many cellular pathways that are critical for genome stability, including DNA repair, chromatin structure, mitosis and apoptosis1. Poly(ADP-ribose) (PAR) is composed of repeating ADP-ribose units linked via a unique glycosidic ribose-ribose bond, and is synthesised from NAD by poly(ADP-ribose) polymerases (PARPs)1,2. Poly(ADP-ribose) glycohydrolase (PARG) is the only protein capable of specific hydrolysis of the ribose-ribose bonds present in PAR chains; its deficiency leads to cell death3,4. Here we show that filamentous fungi and a number of bacteria possess a divergent form of PARG that exhibits all the main characteristics of the human PARG enzyme. We present the first PARG crystal structure (derived from the bacterium Thermomonospora curvata), which reveals that the PARG catalytic domain is a distant member of the ubiquitous ADP-ribose-binding macro domain family5,6. High resolution structures of T. curvata PARG in complexes with ADP-ribose and the PARG inhibitor ADP-HPD, complemented by biochemical studies, allow us to propose a model for PAR binding and catalysis by PARG. Our insights into the PARG structure and catalytic mechanism should greatly improve our understanding of how PARG activity controls reversible protein poly(ADP-ribosyl)ation and potentially of how the defects in this regulation link to human disease.