# Defining synthetic lethal relationships with loss of the homologous recombination factor Rad52

> **NIH NIH F32** · BECKMAN RESEARCH INSTITUTE/CITY OF HOPE · 2023 · $69,500

## Abstract

SUMMARY. The long-term goal of this project is to define factors and pathways that are synthetic lethal with
loss of the human Rad52 protein. Rad52 plays essential roles in several homology-driven DNA repair pathways,
including single strand annealing, transcription-coupled homologous recombination, and mitotic DNA synthesis
(MiDAS). Although Rad52 is not essential, Rad52 loss with disruption of either the breast cancer 1 (BRCA1) or
breast cancer 2 (BRCA2) genes is synthetic lethal. Thus, Rad52 is an intriguing potential target for treatment of
BRCA-deficient cancers. However, the full breadth of pathways and factors that create a state of Rad52-
dependence when compromised are not understood, and the long-term goal of this proposal is to address this
gap in knowledge. In preliminary data, I present my CRISPR knock-out screen in Rad52 Knock-out (Rad52KO)
cells vs. wild-type (Rad52WT) to identify factors that are synthetic lethal with Rad52 (defined here as loss of
fitness). I then present secondary screening that identified three top hits causing increased persistent DNA
damage and loss of viability in the Rad52KO vs Rad52WT: ERCC6L/PICH, DHX9 and GLE1. From these data, my
overall hypothesis is that a key regulator of Mitosis (PICH) and RNA metabolism factors (DHX9 and GLE1) are
synthetic lethal with Rad52 due to dependence on Rad52 to resolve replication stress from diverse sources. Aim
1: To define the synthetic lethal relationship between Rad52 and PICH. Rad52 protects genome stability through
roles in MiDAS and suppression of replication stress. PICH mediates resolution of anaphase ultrafine bridges
(UFBs), a separate pathway to mitigate replication stress. Aim 1a: I posit that these two pathways are partially
redundant in preventing accumulation of genotoxic damage tied to replication stress. Namely, I posit that PICH-
UFBs will be elevated in Rad52KO cells, and conversely that depletion of PICH will cause elevated Rad52
recruitment to replication stress in mitotic cells, as well as MiDAS. Aim 1b: I also posit that replication stress that
persists until mitosis is the source of persistent DNA damage in cells lacking Rad52 and PICH. I will test this by
assaying phosphorylated RPA2 (pRPA), γH2AX, and FANCD2 localization in mitotic cells. Aim 2: To define the
synthetic lethal relationship between Rad52 and RNA metabolism factors DHX9 and GLE1. DHX9 and GLE1
have been shown to suppress RNA-DNA hybrids (R-loops). Thus, I hypothesize that R-loop-related replication
stress underlies synthetic lethality between these genes and Rad52. I will assay whether depletion of these
genes increases R-loops, causes elevated levels of mitotic replication stress (i.e. Rad52 accumulation into foci,
MiDAS, PICH-UFBs, and the measures of replication stress described in Aim 1b). In summary, these studies will
provide insight into how loss of these genes create a dependence on Rad52-mediated mitigation of replication
stress in mitosis (i.e., MiDAS), enhance our under...

## Key facts

- **NIH application ID:** 10678580
- **Project number:** 1F32CA275207-01A1
- **Recipient organization:** BECKMAN RESEARCH INSTITUTE/CITY OF HOPE
- **Principal Investigator:** Beth Anne Osia
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $69,500
- **Award type:** 1
- **Project period:** 2023-04-01 → 2024-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10678580

## Citation

> US National Institutes of Health, RePORTER application 10678580, Defining synthetic lethal relationships with loss of the homologous recombination factor Rad52 (1F32CA275207-01A1). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10678580. Licensed CC0.

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