# Project 3: PARP: PAR-dependent, phase-transitioned protein assemblies and DNA repair

> **NIH NIH P01** · UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB · 2023 · $337,964

## Abstract

SUMMARY- PROJECT 3-PARP: PAR-dependent, phase-transitioned protein assemblies and DNA repair
Poly-ADP-ribose polymerases (PARPs) utilize NAD+ as a donor for ADP-ribose modification of proteins and for
the subsequent additions that generate poly-ADP-ribose (PAR) on these targets. PARP1 and PARP2 are the
predominant enzymes in humans that catalyze PARylation in response to DNA damage and are critical targets
of cancer therapeutics. Rapid formation of PAR occurs at many different types of DNA lesions and has been
shown to be is required for the timely recruitment of DNA repair enzymes. In addition, PAR recruits polypeptides
containing low-complexity domains and intrinsically disordered regions. Some of these disordered proteins form
phase transitions between soluble, gel, and fibrous states. PAR thus seeds phase condensates at sites of DNA
damage to create a specialized compartment for repairing DNA. Despite the fundamental importance of this
phenomenon, we know very little about the structural basis for PARP generation of PAR polymers at DNA
damage sites or the mechanistic role of PAR-seeded phase transitions in promoting DNA repair. Here we
hypothesize that PAR-driven phase separation regulates DNA repair outcomes at diverse DNA lesions and that
the length and structure of PAR polymer is critical for tuning these outcomes. We will test this hypothesis by
investigating the mechanistic basis of ADP-ribose addition and PAR formation by PARP1/2 in the context of
cofactor HPF1 using structural biology, ensemble biochemistry, and single-molecule methods (Aim 1). Structural
insights from this effort will produce a working model for PARP1/2 catalytic activity as well as a toolbox of
PARP1/2 mutants that generate varying lengths and structures of PAR. As part of this effort we will also develop
inhibitors for the primary glycohydrolase that removes DNA damage-induced ADP-ribose modifications (ARH3).
We will probe the role of phase transitions in DNA repair by using FUS and other low-complexity proteins to
investigate the structural basis of their interactions with PAR-seeded domains of various lengths (Aim 2). Lastly,
we will determine the effects of phase transitions on the recruitment and activity of DNA double-strand break
repair complexes in vitro, as well as the effects of these transitions on the repair of multiple types of DNA damage
in human cells (Aim 3). These efforts join P3 with the other projects in SBDR5 by collaborating to identify effects
of lesion-specific PARylation on the repair of alkylation damage, double-strand breaks, and stalled replication
forks. In addition, we will determine the composition of PAR-seeded domains in response to DNA damage and
how this changes in response to clinically used PARP and glycohydrolase inhibitors. This project will produce
structural and mechanistic insights into the dynamic regulation of PAR synthesis and disassembly that occurs in
human cells and will determine how the PAR-seeded phase transitions at ...

## Key facts

- **NIH application ID:** 10695047
- **Project number:** 5P01CA092584-23
- **Recipient organization:** UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
- **Principal Investigator:** TANYA T PAULL
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $337,964
- **Award type:** 5
- **Project period:** 2001-09-27 → 2026-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10695047

## Citation

> US National Institutes of Health, RePORTER application 10695047, Project 3: PARP: PAR-dependent, phase-transitioned protein assemblies and DNA repair (5P01CA092584-23). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10695047. Licensed CC0.

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