Deletion of the 3’RR cis-enhancer IgH element affects DNA repair junctions during B-cell class switch recombination.

Cernunnos is involved in the nonhomologous end-joining (NHEJ) process during DNA double-strand break (DSB) repair. Here, we studied immunoglobulin (Ig) class switch recombination (CSR), a physiological process which relies on proper repair of the DSBs, in B cells from Cernunnos-deficient patients. The pattern of in vivo generated CSR junctions is altered in these cells, with unusually long microhomologies and a lack of direct end-joining. The CSR junctions from Cernunnos-deficient patients largely resemble those from patients lacking DNA ligase IV, Artemis, or ATM, suggesting that these factors are involved in the same end-joining pathway during CSR. By screening 269 mature B cell lymphoma biopsies, we also identified a somatic missense Cernunnos mutation in a diffuse large B cell lymphoma sample. This mutation has a dominant-negative effect on joining of a subset of DNA ends in an in vitro NHEJ assay. Translocations involving both Ig heavy chain loci and clonal-like, dynamic IgA switching activities were observed in this tumor. Collectively, our results suggest a link between defects in the Cernunnos-dependent NHEJ pathway and aberrant CSR or switch translocations during the development of B cell malignancies.

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MicroRNA-directed pathway discovery elucidates an miR-221/222–mediated regulatory circuit in class switch recombination

Journal of Experimental Medicine ◽

10.1084/jem.20201422 ◽

2021 ◽

Vol 218 (11) ◽

Author(s):

Eric J. Wigton ◽

Yohei Mikami ◽

Ryan J. McMonigle ◽

Carlos A. Castellanos ◽

Adam K. Wade-Vallance ◽

...

Keyword(s):

B Cells ◽

B Cell ◽

Cell Fate ◽

Cell Activation ◽

Class Switch Recombination ◽

Cell Fate Decisions ◽

Class Switch ◽

Mrna Targets ◽

Regulatory Circuit ◽

Pathway Discovery

MicroRNAs (miRNAs, miRs) regulate cell fate decisions by post-transcriptionally tuning networks of mRNA targets. We used miRNA-directed pathway discovery to reveal a regulatory circuit that influences Ig class switch recombination (CSR). We developed a system to deplete mature, activated B cells of miRNAs, and performed a rescue screen that identified the miR-221/222 family as a positive regulator of CSR. Endogenous miR-221/222 regulated B cell CSR to IgE and IgG1 in vitro, and miR-221/222–deficient mice exhibited defective IgE production in allergic airway challenge and polyclonal B cell activation models in vivo. We combined comparative Ago2-HITS-CLIP and gene expression analyses to identify mRNAs bound and regulated by miR-221/222 in primary B cells. Interrogation of these putative direct targets uncovered functionally relevant downstream genes. Genetic depletion or pharmacological inhibition of Foxp1 and Arid1a confirmed their roles as key modulators of CSR to IgE and IgG1.

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Phf5a regulates DNA repair in class switch recombination via p400 and histone H2A variant deposition

The EMBO Journal ◽

10.15252/embj.2020106393 ◽

2021 ◽

Author(s):

Nasim A Begum ◽

Farazul Haque ◽

Andre Stanlie ◽

Afzal Husain ◽

Samiran Mondal ◽

...

Keyword(s):

Dna Repair ◽

Class Switch Recombination ◽

Histone H2a ◽

Class Switch

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Clinical, Immunological, and Functional Characterization of Six Patients with Very High IgM Levels

Journal of Clinical Medicine ◽

10.3390/jcm9030818 ◽

2020 ◽

Vol 9 (3) ◽

pp. 818 ◽

Cited By ~ 1

Author(s):

Vera Gallo ◽

Emilia Cirillo ◽

Rosaria Prencipe ◽

Alessio Lepore ◽

Luigi Del Vecchio ◽

...

Keyword(s):

B Cell ◽

Somatic Hypermutation ◽

Functional Characterization ◽

Class Switch Recombination ◽

Pathogenic Mutation ◽

Potential Candidate ◽

Class Switch ◽

Potential Candidate Gene ◽

Very High

Very high IgM levels represent the hallmark of hyper IgM (HIGM) syndromes, a group of primary immunodeficiencies (PIDs) characterized by susceptibility to infections and malignancies. Other PIDs not fulfilling the diagnostic criteria for HIGM syndromes can also be characterized by high IgM levels and susceptibility to malignancies. The aim of this study is to characterize clinical phenotype, immune impairment, and pathogenic mechanism in six patients with very high IgM levels in whom classical HIGM syndromes were ruled out. The immunological analysis included extended B-cell immunophenotyping, evaluation of class switch recombination and somatic hypermutation, and next generation sequencing (NGS). Recurrent or severe infections and chronic lung changes at the diagnosis were reported in five out of six and two out of six patients, respectively. Five out of six patients showed signs of lymphoproliferation and four patients developed malignancies. Four patients showed impaired B-cell homeostasis. Class switch recombination was functional in vivo in all patients. NGS revealed, in one case, a pathogenic mutation in PIK3R1. In a second case, the ITPKB gene, implicated in B- and T-cell development, survival, and activity was identified as a potential candidate gene. Independent of the genetic basis, very high IgM levels represent a risk factor for the development of recurrent infections leading to chronic lung changes, lymphoproliferation, and high risk of malignancies.

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Lsh/HELLS is required for B lymphocyte development and immunoglobulin class switch recombination

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2004112117 ◽

2020 ◽

Vol 117 (33) ◽

pp. 20100-20108

Author(s):

Yafeng He ◽

Jianke Ren ◽

Xiaoping Xu ◽

Kai Ni ◽

Andrew Schwader ◽

...

Keyword(s):

Bone Marrow ◽

Bone Marrow Transplantation ◽

B Cell ◽

Marrow Transplantation ◽

Class Switch Recombination ◽

B Cell Development ◽

Hematopoietic Cell ◽

Cell Numbers ◽

Class Switch

Mutation of HELLS (Helicase, Lymphoid-Specific)/Lsh in human DNA causes a severe immunodeficiency syndrome, but the nature of the defect remains unknown. We assessed here the role of Lsh in hematopoiesis using conditional Lsh knockout mice with expression of Mx1 or Vav Cre-recombinase. Bone marrow transplantation studies revealed that Lsh depletion in hematopoietic stem cells severely reduced B cell numbers and impaired B cell development in a hematopoietic cell-autonomous manner. Lsh-deficient mice without bone marrow transplantation exhibited lower Ig levels in vivo compared to controls despite normal peripheral B cell numbers. Purified B lymphocytes proliferated normally but produced less immunoglobulins in response to in vitro stimulation, indicating a reduced capacity to undergo class switch recombination (CSR). Analysis of germline transcripts, examination of double-stranded breaks using biotin-labeling DNA break assay, and End-seq analysis indicated that the initiation of the recombination process was unscathed. In contrast, digestion–circularization PCR analysis and high-throughput sequencing analyses of CSR junctions and a chromosomal break repair assay indicated an impaired ability of the canonical end-joining pathway in Lsh-deficient B cells. Our data suggest a hematopoietic cell-intrinsic role of Lsh in B cell development and in CSR providing a potential target for immunodeficiency therapy.

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