# Homologous Recombination Repair Domains: Formation and Impact on Genome Stability

> **NIH NIH F30** · COLUMBIA UNIVERSITY HEALTH SCIENCES · 2022 · $51,752

## Abstract

PROJECT SUMMARY: DNA double-strand break (DSB) repair is spatially organized into nuclear repair domains
that specifically facilitate DSB repair by homologous recombination (HR). HR, one of the major DSB pathways
along with non-homologous end-joining, has been implicated in tumorigenesis, notably following mutations in
the tumor suppressor genes BRCA1 and BRCA2 [1, 2]. Our lab demonstrated that upon DSB formation by
induction of a restriction endonuclease (RE) or treatment with neocarzinostatin (NCS), WASP activates ARP2/3,
which polymerizes nuclear actin into branched filaments [4]. This enhances the mobility of DSBs destined for HR
and their subsequent clustering into HR domains.
 The DNA topoisomerase II (Top2) inhibitor etoposide (ETO) yields DSBs harboring protein-DNA adducts that
require resection and subsequent repair by HR factors, including MRN, CtIP, and BRCA1 [5, 6]. Because of the
absolute requirement for poisoned Top2 removal prior to repair, ETO is a unique way to probe the functional
relationship between resection and movement. ETO is used to treat a wide range of cancers, including leukemia
and soft tissue cancers. However, treatment is associated with secondary leukemias due to translocations. Using
live-cell imaging, I show that ETO DSBs undergo ARP2/3-mediated movement and clustering. However, unlike
RE and NCS DSBs, movement is not restricted to G2 but also occurs in G1. Additionally, ETO breaks in G1
undergo resection and load HR machinery, such as RPA. I have also begun examining the role of HR factors,
including Mre11 and BRCA2, in repair domain formation following the generation of DSBs by RE, NCS and ETO.
 Although DSB clustering is crucial for HR, little is known about how repair domains are formed and their local
and genome-wide implications. For example, we do not fully understand the crosstalk between movement (actin,
WASP) and repair (HR machinery) in mammalian cells. Additionally, the dynamics of DSBs likely influences
chromosomal rearrangements. Our lab is integrating high-throughput genomic technologies that assess gene-
gene interactions and translocation events to determine the genome-wide implications of DSB mobility. The
overarching goals of this study are to elucidate mechanisms by which nuclear actin polymerization and HR
proteins regulate repair domain formation and to evaluate the genome-wide impact of DSB mobility. I
hypothesize that HR proteins, including the resection machinery, play a critical role in regulating ARP2/3-
mediated DSB movements and subsequent clustering. I further propose that nuclear actin polymerization
impacts genome organization following DNA damage and thus affects translocation frequency. I will investigate
these hypotheses in the following aims:
Aim 1: Elucidate the contribution of HR machinery to Arp2/3-dependent DSB clustering.
Aim 2: Determine the impact of ARP2/3-mediated DSB movement on genome stability.

## Key facts

- **NIH application ID:** 10440346
- **Project number:** 5F30CA250166-03
- **Recipient organization:** COLUMBIA UNIVERSITY HEALTH SCIENCES
- **Principal Investigator:** Jennifer Ashley Zagelbaum
- **Activity code:** F30 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $51,752
- **Award type:** 5
- **Project period:** 2020-07-01 → 2023-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10440346

## Citation

> US National Institutes of Health, RePORTER application 10440346, Homologous Recombination Repair Domains: Formation and Impact on Genome Stability (5F30CA250166-03). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10440346. Licensed CC0.

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