# Defining the Mechanism and Function of DNA Double Strand Break Induced Inhibition of V(D)J Recombination

> **NIH NIH F31** · UNIVERSITY OF PENNSYLVANIA · 2021 · $46,036

## Abstract

Project Summary: B cell-mediated adaptive immunity relies on the programmed induction of DNA double strand
breaks (DSBs) to create diverse immunoglobin (Ig) gene repertoires. The RAG1/RAG2 (RAG) endonuclease
assembles Ig genes through recombination of variable (V), diversity (D), and joining (J) gene segments of Ig loci.
This process is vital for adaptive immunity and mammalian survival yet, it also confers risk, as most B lineage
cancers contain clonal translocations involving an Ig locus and a proto-oncogene, underscoring the vital
importance of tightly regulating V(D)J recombination. My thesis lab discovered that RAG or genotoxic DSBs
rapidly repress transcription of the Rag1/2 locus by signaling via the ATM kinase, a key regulator of the cellular
DSB response. Atm-/- mice have higher frequencies of developing B cells with RAG DSBs at both Ig alleles and
of mature B lymphocytes with Ig translocations. Moreover, B cell-specific Atm deletion in mice increases the
incidence of B lineage lymphomas with Ig translocations. These data are consistent with DSB-induced repression
of Rag1/2 being critical to protect from Ig translocations; yet, given the multifunctional roles of ATM in the DSB
response, these phenotypes cannot be directly attributed to DSB-induced repression of Rag1/2 expression.
Genotoxic DSBs induce ATM-dependent phosphorylation of the NFκB essential modulator (Nemo) protein to
activate NFκB transcription factors. I show that Nemo-/- developing B cells have impaired repression of Rag1/2
in response to DSBs and display increased RAG DSBs at Ig loci. My data are consistent with prior Atm-/- studies,
but unlike Atm-/- cells, Nemo-/- cells retain normal DSB repair and checkpoint/apoptosis activation. Thus, the
Nemo-/- model provides me an opportunity to more directly test my central hypothesis that DSBs induce
Nemo/NFκB-mediated transcriptional repression of Rag1/2 to suppress Ig translocations and resultant lymphoid
cancers. I propose to elucidate the mechanism of Nemo-dependent Rag1/2 repression. Aim 1 of this proposal
will test the hypothesis that NFκB factors directly bind the Rag1/2 Erag enhancer to mediate DSB-induced
repression of Rag1/2 by inhibiting transcriptional elongation. This may reveal novel mechanisms of NFκB-
mediated transcriptional repression, which are largely undefined. I will determine DSB-induced NFκB binding
sites and their functional role in Rag1/2 repression. Furthermore, I will define the transcriptional state of Rag1/2
in the absence or presence of DSBs to determine the mechanistic levels at which DSBs repress Rag1/2. Aim 2
will test the hypothesis that DSB-induced repression of Rag1/2 limits RAG DSBs and thereby suppresses
oncogenic Ig translocations. Mice with B cell specific deletion of Nemo will be used to i) quantify RAG DSBs at
Ig alleles in pro-B and pre-B cells, ii) quantify Ig translocations in non-malignant B lineage cells, and iii) evaluate
predisposition to B lineage cancers with oncogenic Ig transl...

## Key facts

- **NIH application ID:** 10139829
- **Project number:** 1F31AI152354-01A1
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **Principal Investigator:** Rebecca Ann Glynn
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $46,036
- **Award type:** 1
- **Project period:** 2021-01-01 → 2023-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10139829

## Citation

> US National Institutes of Health, RePORTER application 10139829, Defining the Mechanism and Function of DNA Double Strand Break Induced Inhibition of V(D)J Recombination (1F31AI152354-01A1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10139829. Licensed CC0.

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