Robust immunoglobulin class switch recombination and end joining inParp9-deficient mice

DNA double-strand breaks (DSBs) introduced in the switch (S) regions are intermediates during immunoglobulin class switch recombination (CSR). These breaks are subsequently recognized, processed, and joined, leading to recombination of the two S regions. Nonhomologous end-joining (NHEJ) is believed to be the principle mechanism involved in DSB repair during CSR. One important component in NHEJ, Artemis, has however been considered to be dispensable for efficient CSR. In this study, we have characterized the S recombinational junctions from Artemis-deficient human B cells. Sμ–Sα junctions could be amplified from all patients tested and were characterized by a complete lack of “direct” end-joining and a remarkable shift in the use of an alternative, microhomology-based end-joining pathway. Sμ–Sγ junctions could only be amplified from one patient who carries “hypomorphic” mutations. Although these Sμ–Sγ junctions appear to be normal, a significant increase of an unusual type of sequential switching from immunoglobulin (Ig)M, through one IgG subclass, to a different IgG subclass was observed, and the Sγ–Sγ junctions showed long microhomologies. Thus, when the function of Artemis is impaired, varying modes of CSR junction resolution may be used for different S regions. Our findings strongly link Artemis to the predominant NHEJ pathway during CSR.

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Parp1 facilitates alternative NHEJ, whereas Parp2 suppresses IgH/c-myc translocations during immunoglobulin class switch recombination

Journal of Experimental Medicine ◽

10.1084/jem.20082468 ◽

2009 ◽

Vol 206 (5) ◽

pp. 1047-1056 ◽

Cited By ~ 121

Author(s):

Isabelle Robert ◽

Françoise Dantzer ◽

Bernardo Reina-San-Martin

Keyword(s):

Dna Damage ◽

Cytidine Deaminase ◽

Class Switch Recombination ◽

Dependent Manner ◽

Dna Breaks ◽

End Joining ◽

Immunoglobulin Class ◽

Class Switch ◽

Switch Regions ◽

Alternative Nhej

Immunoglobulin class switch recombination (CSR) is initiated by DNA breaks triggered by activation-induced cytidine deaminase (AID). These breaks activate DNA damage response proteins to promote appropriate repair and long-range recombination. Aberrant processing of these breaks, however, results in decreased CSR and/or increased frequency of illegitimate recombination between the immunoglobulin heavy chain locus and oncogenes like c-myc. Here, we have examined the contribution of the DNA damage sensors Parp1 and Parp2 in the resolution of AID-induced DNA breaks during CSR. We find that although Parp enzymatic activity is induced in an AID-dependent manner during CSR, neither Parp1 nor Parp2 are required for CSR. We find however, that Parp1 favors repair of switch regions through a microhomology-mediated pathway and that Parp2 actively suppresses IgH/c-myc translocations. Thus, we define Parp1 as facilitating alternative end-joining and Parp2 as a novel translocation suppressor during CSR.

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Regulation of DNA End Joining, Resection, and Immunoglobulin Class Switch Recombination by 53BP1

Molecular Cell ◽

10.1016/j.molcel.2011.03.019 ◽

2011 ◽

Vol 42 (3) ◽

pp. 319-329 ◽

Cited By ~ 152

Author(s):

Anne Bothmer ◽

Davide F. Robbiani ◽

Michela Di Virgilio ◽

Samuel F. Bunting ◽

Isaac A. Klein ◽

...

Keyword(s):

Class Switch Recombination ◽

End Joining ◽

Immunoglobulin Class ◽

Class Switch

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Faculty Opinions recommendation of Regulation of DNA end joining, resection, and immunoglobulin class switch recombination by 53BP1.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.12764956.14036059 ◽

2011 ◽

Author(s):

Penny Jeggo ◽

Aaron Goodarzi

Keyword(s):

Class Switch Recombination ◽

End Joining ◽

Immunoglobulin Class ◽

Class Switch

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Elucidation of the enigmatic IgD class-switch recombination via germline deletion of the IgH 3′ regulatory region

Journal of Experimental Medicine ◽

10.1084/jem.20131385 ◽

2014 ◽

Vol 211 (5) ◽

pp. 975-985 ◽

Cited By ~ 41

Author(s):

Pauline Rouaud ◽

Alexis Saintamand ◽

Faten Saad ◽

Claire Carrion ◽

Sandrine Lecardeur ◽

...

Keyword(s):

Somatic Hypermutation ◽

Regulatory Region ◽

Class Switch Recombination ◽

End Joining ◽

Class Switch ◽

Classical Class ◽

Mesenteric Lymph ◽

Alternative End Joining ◽

Activation Induced Deaminase ◽

Deficient Mice

Classical class-switch recombination (cCSR) substitutes the Cμ gene with Cγ, Cε, or Cα, thereby generating IgG, IgE, or IgA classes, respectively. This activation-induced deaminase (AID)–driven process is controlled by the IgH 3′ regulatory region (3′RR). Regulation of rare IgD CSR events has been enigmatic. We show that μδCSR occurs in mouse mesenteric lymph node (MLN) B cells and is AID-dependent. AID attacks differ from those in cCSR because they are not accompanied by extensive somatic hypermutation (SHM) of targeted regions and because repaired junctions exhibit features of the alternative end-joining (A-EJ) pathway. In contrast to cCSR and SHM, μδCSR is 3′RR-independent, as its absence affects neither breakpoint locations in Sμ- and Sδ-like (σδ) nor mutation patterns at Sμ-σδ junctions. Although mutations occur in the immediate proximity of the μδ junctions, SHM is absent distal to the junctions within both Sμ and rearranged VDJ regions. In conclusion, μδCSR is active in MLNs, occurs independently of 3′RR-driven assembly, and is even dramatically increased in 3′RR-deficient mice, further showing that its regulation differs from cCSR.

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Immunoglobulin Class Switch Recombination Is Impaired in Atm-deficient Mice

Journal of Experimental Medicine ◽

10.1084/jem.20041074 ◽

2004 ◽

Vol 200 (9) ◽

pp. 1111-1121 ◽

Cited By ~ 118

Author(s):

Joanne M. Lumsden ◽

Thomas McCarty ◽

Lisa K. Petiniot ◽

Rhuna Shen ◽

Carrolee Barlow ◽

...

Keyword(s):

B Cells ◽

Dna Recombination ◽

Class Switch Recombination ◽

Intrinsic Defect ◽

Wild Type ◽

Immunoglobulin Class ◽

Class Switch ◽

Deficient Mice

Immunoglobulin class switch recombination (Ig CSR) involves DNA double strand breaks (DSBs) at recombining switch regions and repair of these breaks by nonhomologous end-joining. Because the protein kinase ataxia telengiectasia (AT) mutated (ATM) plays a critical role in DSB repair and AT patients show abnormalities of Ig isotype expression, we assessed the role of ATM in CSR by examining ATM-deficient mice. In response to T cell–dependent antigen (Ag), Atm−/− mice secreted substantially less Ag-specific IgA, IgG1, IgG2b, and IgG3, and less total IgE than Atm+/+ controls. To determine whether Atm−/− B cells have an intrinsic defect in their ability to undergo CSR, we analyzed in vitro responses of purified B cells. Atm−/− cells secreted substantially less IgA, IgG1, IgG2a, IgG3, and IgE than wild-type (WT) controls in response to stimulation with lipopolysaccharide, CD40 ligand, or anti-IgD plus appropriate cytokines. Molecular analysis of in vitro responses indicated that WT and Atm−/− B cells produced equivalent amounts of germline IgG1 and IgE transcripts, whereas Atm−/− B cells produced markedly reduced productive IgG1 and IgE transcripts. The reduction in isotype switching by Atm−/− B cells occurs at the level of genomic DNA recombination as measured by digestion–circularization PCR. Analysis of sequences at CSR sites indicated that there is greater microhomology at the μ–γ1 switch junctions in ATM B cells than in wild-type B cells, suggesting that ATM function affects the need or preference for sequence homology in the CSR process. These findings suggest a role of ATM in DNA DSB recognition and/or repair during CSR.

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Aberrant recombination and repair during immunoglobulin class switching in BRCA1-deficient human B cells

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1418947112 ◽

2015 ◽

Vol 112 (7) ◽

pp. 2157-2162 ◽

Cited By ~ 9

Author(s):

Andrea Björkman ◽

Per Qvist ◽

Likun Du ◽

Margarita Bartish ◽

Apostolos Zaravinos ◽

...

Keyword(s):

Breast Cancer ◽

B Cells ◽

Class Switch Recombination ◽

Cancer Type ◽

End Joining ◽

Immunoglobulin Class ◽

Class Switch ◽

Human B Cells ◽

Breast Cancer Type

Breast cancer type 1 susceptibility protein (BRCA1) has a multitude of functions that contribute to genome integrity and tumor suppression. Its participation in the repair of DNA double-strand breaks (DSBs) during homologous recombination (HR) is well recognized, whereas its involvement in the second major DSB repair pathway, nonhomologous end-joining (NHEJ), remains controversial. Here we have studied the role of BRCA1 in the repair of DSBs in switch (S) regions during immunoglobulin class switch recombination, a physiological, deletion/recombination process that relies on the classical NHEJ machinery. A shift to the use of microhomology-based, alternative end-joining (A-EJ) and increased frequencies of intra-S region deletions as well as insertions of inverted S sequences were observed at the recombination junctions amplified from BRCA1-deficient human B cells. Furthermore, increased use of long microhomologies was found at recombination junctions derived from E3 ubiquitin-protein ligase RNF168-deficient, Fanconi anemia group J protein (FACJ, BRIP1)-deficient, or DNA endonuclease RBBP8 (CtIP)-compromised cells, whereas an increased frequency of S-region inversions was observed in breast cancer type 2 susceptibility protein (BRCA2)-deficient cells. Thus, BRCA1, together with its interaction partners, seems to play an important role in repairing DSBs generated during class switch recombination by promoting the classical NHEJ pathway. This may not only provide a general mechanism underlying BRCA1’s function in maintaining genome stability and tumor suppression but may also point to a previously unrecognized role of BRCA1 in B-cell lymphomagenesis.

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Impaired induction of DNA lesions during immunoglobulin class-switch recombination in humans influences end-joining repair

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1012591108 ◽

2010 ◽

Vol 107 (51) ◽

pp. 22225-22230 ◽

Cited By ~ 27

Author(s):

S. Kracker ◽

K. Imai ◽

P. Gardes ◽

H. D. Ochs ◽

A. Fischer ◽

...

Keyword(s):

Class Switch Recombination ◽

Dna Lesions ◽

End Joining ◽

Immunoglobulin Class ◽

Class Switch

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A regulatory role for the cohesin loader NIPBL in nonhomologous end joining during immunoglobulin class switch recombination

Journal of Experimental Medicine ◽

10.1084/jem.20130168 ◽

2013 ◽

Vol 210 (12) ◽

pp. 2503-2513 ◽

Cited By ~ 28

Author(s):

Elin Enervald ◽

Likun Du ◽

Torkild Visnes ◽

Andrea Björkman ◽

Emma Lindgren ◽

...

Keyword(s):

Class Switch Recombination ◽

Nonhomologous End Joining ◽

Loss Of Function ◽

Dna Double Strand Breaks ◽

End Joining ◽

Gene Encoding ◽

Immunoglobulin Class ◽

Cellular Processes ◽

Class Switch ◽

Cornelia De Lange

DNA double strand breaks (DSBs) are mainly repaired via homologous recombination (HR) or nonhomologous end joining (NHEJ). These breaks pose severe threats to genome integrity but can also be necessary intermediates of normal cellular processes such as immunoglobulin class switch recombination (CSR). During CSR, DSBs are produced in the G1 phase of the cell cycle and are repaired by the classical NHEJ machinery. By studying B lymphocytes derived from patients with Cornelia de Lange Syndrome, we observed a strong correlation between heterozygous loss-of-function mutations in the gene encoding the cohesin loading protein NIPBL and a shift toward the use of an alternative, microhomology-based end joining during CSR. Furthermore, the early recruitment of 53BP1 to DSBs was reduced in the NIPBL-deficient patient cells. Association of NIPBL deficiency and impaired NHEJ was also observed in a plasmid-based end-joining assay and a yeast model system. Our results suggest that NIPBL plays an important and evolutionarily conserved role in NHEJ, in addition to its canonical function in sister chromatid cohesion and its recently suggested function in HR.

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