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IgH class switch recombination to IgG1 in DNA-PKcs-deficient B cells
John P Manis1, Darryll Dudley, Lianne Kaylor
1Howard Hughes Medical Institute and Children's Hospital, Center for Blood Research and Department of Genetics, Harvard Medical School, 300 Longwood Avenue, Boston, MA 02115, USA.
Insights
DNA-PKcs protein is essential for most immunoglobulin heavy chain class switch recombination (CSR) but not for IgG1 switching. This DNA-PKcs-independent IgG1 CSR mechanism is a key finding.
Area of Science:
- Immunology
- Molecular Biology
- Genetics
Background:
- Immunoglobulin heavy chain class switch recombination (CSR) is a critical adaptive immune process.
- Nonhomologous DNA end-joining (NHEJ) pathway proteins, including DNA-PKcs, are implicated in DNA repair and V(D)J recombination.
- The specific role of DNA-PKcs in CSR remains incompletely understood.
Purpose of the Study:
- To investigate the role of DNA-PKcs in immunoglobulin heavy chain class switch recombination (CSR).
- To determine if DNA-PKcs deficiency affects B cell activation, proliferation, and germline gene transcription.
Main Methods:
- Generation of DNA-PKcs-deficient B cells (DP-T) through complementation of DP-T mice with Ig heavy and light chain knock-in transgenes (DP-T/HC/LC mice).
- Assay of CSR ability in DP-T/HC/LC B cells upon stimulation.
- Analysis of B cell proliferation, germline C(H) gene transcription, and AID induction.
Main Results:
- DP-T/HC/LC mice exhibited severe deficiency in most serum IgH isotypes, except IgM and unexpectedly, IgG1.
- DNA-PKcs-deficient B cells showed normal proliferation, germline C(H) gene transcription, and AID induction.
- In vitro activated DP-T/HC/LC B cells exclusively underwent switching to IgG1, with frequent switching to gamma1 on both chromosomes.
Conclusions:
- DNA-PKcs is essential for CSR to the majority of C(H) genes.
- Class switch recombination to IgG1 (gamma1) proceeds through a DNA-PKcs-independent mechanism.
- This study reveals a novel DNA-PKcs-independent pathway for IgG1 switching.
Abstract:
To assess the role of the DNA-PKcs nonhomologous DNA end-joining (NHEJ) protein in Ig heavy chain class switch recombination (CSR), we assayed CSR ability of DNA-PKcs-deficient (DP-T) B cells generated via complementation of DP-T mice with Ig heavy chain and light chain knock-in transgenes (DP-T/HC/LC mice). DP-T/HC/LC mice were severely deficient for all serum IgH isotypes except IgM and, unexpectedly, IgG1. Upon appropriate stimulation, DP-T/HC/LC B cells showed normal proliferation, germline C(H) gene transcription, and AID induction, indicating that DNA-PKcs deficiency did not affect cellular events upstream to CSR. Yet, in vitro activated DP-T/HC/LC B cells again underwent switching only to IgG1 and, like wild-type cells, frequently underwent CSR to gamma1 on both chromosomes. We conclude that DNA-PKcs is required for CSR to most C(H) genes but that CSR to gamma1 occurs via a DNA-PKcs-independent mechanism.