scholarly journals Fixing DNA breaks during class switch recombination

2008 ◽  
Vol 205 (3) ◽  
pp. 509-513 ◽  
Author(s):  
Christopher J. Jolly ◽  
Adam J.L. Cook ◽  
John P. Manis
Keyword(s):  
Dna Sequences ◽  
Class Switch Recombination ◽  
Nonhomologous End Joining ◽  
Repair Pathway ◽  
Dna Breaks ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Class Switch ◽  
Dependent Protein

Immunoglobulin (Ig) class switch recombination (CSR) involves the breakage and subsequent repair of two DNA sequences, known as switch (S) regions, which flank IgH constant region exons. The resolution of CSR-associated breaks is thought to require the nonhomologous end-joining (NHEJ) DNA repair pathway, but the role of the NHEJ factor DNA-dependent protein kinase catalytic subunit (DNA-PKcs) in this process has been unclear. A new study, in which broken IgH-containing chromosomes in switching B cells were visualized directly, clearly demonstrated that DNA-PKcs and, unexpectedly, the nuclease Artemis are involved in the resolution of switch breaks.

scholarly journals The cohesin complex regulates immunoglobulin class switch recombination

2013 ◽  
Vol 210 (12) ◽  
pp. 2495-2502 ◽  
Author(s):  
Anne-Sophie Thomas-Claudepierre ◽  
Ebe Schiavo ◽  
Vincent Heyer ◽  
Marjorie Fournier ◽  
Adeline Page ◽  
...  
Keyword(s):  
Cytidine Deaminase ◽  
Class Switch Recombination ◽  
Nonhomologous End Joining ◽  
Cohesin Complex ◽  
Dna Breaks ◽  
End Joining ◽  
Regulatory Subunits ◽  
Class Switch ◽  
Chromatid Cohesion ◽  
Switch Regions

Immunoglobulin (Ig) class switch recombination (CSR) is initiated by the transcription-coupled recruitment of activation-induced cytidine deaminase (AID) to switch regions and by the subsequent generation of double-stranded DNA breaks (DSBs). These DNA breaks are ultimately resolved through the nonhomologous end joining (NHEJ) pathway. We show that during CSR, AID associates with subunits of cohesin, a complex previously implicated in sister chromatid cohesion, DNA repair, and the formation of DNA loops between enhancers and promoters. Furthermore, we implicate the cohesin complex in the mechanism of CSR by showing that cohesin is dynamically recruited to the Sμ-Cμ region of the IgH locus during CSR and that knockdown of cohesin or its regulatory subunits results in impaired CSR and increased usage of microhomology-based end joining.

scholarly journals Loss of ZBTB24 impairs nonhomologous end-joining and class-switch recombination in patients with ICF syndrome

2020 ◽  
Vol 217 (11) ◽  
Author(s):  
Angela Helfricht ◽  
Peter E. Thijssen ◽  
Magdalena B. Rother ◽  
Rashmi G. Shah ◽  
Likun Du ◽  
...  
Keyword(s):  
Autosomal Recessive ◽  
Class Switch Recombination ◽  
Nonhomologous End Joining ◽  
Dna Breaks ◽  
End Joining ◽  
Immunoglobulin Production ◽  
Icf Syndrome ◽  
Class Switch ◽  
Centromeric Instability ◽  
Gene Defects

The autosomal recessive immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome is a genetically heterogeneous disorder. Despite the identification of the underlying gene defects, it is unclear how mutations in any of the four known ICF genes cause a primary immunodeficiency. Here we demonstrate that loss of ZBTB24 in B cells from mice and ICF2 patients affects nonhomologous end-joining (NHEJ) during immunoglobulin class-switch recombination and consequently impairs immunoglobulin production and isotype balance. Mechanistically, we found that ZBTB24 associates with poly(ADP-ribose) polymerase 1 (PARP1) and stimulates its auto-poly(ADP-ribosyl)ation. The zinc-finger in ZBTB24 binds PARP1-associated poly(ADP-ribose) chains and mediates the PARP1-dependent recruitment of ZBTB24 to DNA breaks. Moreover, through its association with poly(ADP-ribose) chains, ZBTB24 protects them from degradation by poly(ADP-ribose) glycohydrolase (PARG). This facilitates the poly(ADP-ribose)-dependent assembly of the LIG4/XRCC4 complex at DNA breaks, thereby promoting error-free NHEJ. Thus, we uncover ZBTB24 as a regulator of PARP1-dependent NHEJ and class-switch recombination, providing a molecular basis for the immunodeficiency in ICF2 syndrome.

scholarly journals Kinase-dependent structural role of DNA-PKcs during immunoglobulin class switch recombination

2018 ◽  
Vol 115 (34) ◽  
pp. 8615-8620 ◽  
Author(s):  
Jennifer L. Crowe ◽  
Zhengping Shao ◽  
Xiaobin S. Wang ◽  
Pei-Chi Wei ◽  
Wenxia Jiang ◽  
...  
Keyword(s):  
B Cells ◽  
High Throughput Sequencing ◽  
Class Switch Recombination ◽  
End Joining ◽  
Extensive Resection ◽  
Structural Role ◽  
Class Switch ◽  
Cell Class ◽  
Dependent Protein

The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is a classical nonhomologous end-joining (cNHEJ) factor. Loss of DNA-PKcs diminished mature B cell class switch recombination (CSR) to other isotypes, but not IgG1. Here, we show that expression of the kinase-dead DNA-PKcs (DNA-PKcsKD/KD) severely compromises CSR to IgG1. High-throughput sequencing analyses of CSR junctions reveal frequent accumulation of nonproductive interchromosomal translocations, inversions, and extensive end resection in DNA-PKcsKD/KD, but not DNA-PKcs−/−, B cells. Meanwhile, the residual joints from DNA-PKcsKD/KD cells and the efficient Sµ-Sγ1 junctions from DNA-PKcs−/− B cells both display similar preferences for small (2–6 nt) microhomologies (MH). In DNA-PKcs−/− cells, Sµ-Sγ1 joints are more resistant to inversions and extensive resection than Sµ-Sε and Sµ-Sµ joints, providing a mechanism for the isotype-specific CSR defects. Together, our findings identify a kinase-dependent role of DNA-PKcs in suppressing MH-mediated end joining and a structural role of DNA-PKcs protein in the orientation of CSR.

scholarly journals DNA-dependent Protein Kinase Activity Is Not Required for Immunoglobulin Class Switching

2002 ◽  
Vol 196 (11) ◽  
pp. 1483-1495 ◽  
Author(s):  
Gayle C. Bosma ◽  
Jiyoon Kim ◽  
Teresa Urich ◽  
Donna M. Fath ◽  
Maria G. Cotticelli ◽  
...  
Keyword(s):  
B Cells ◽  
Protein Kinase ◽  
Scid Mice ◽  
Nonhomologous End Joining ◽  
Protein Kinase Activity ◽  
Dna Double Strand Breaks ◽  
Dna Breaks ◽  
End Joining ◽  
Dependent Protein Kinase ◽  
Dependent Protein

Class switch recombination (CSR), similar to V(D)J recombination, is thought to involve DNA double strand breaks and repair by the nonhomologous end–joining pathway. A key component of this pathway is DNA-dependent protein kinase (DNA-PK), consisting of a catalytic subunit (DNA-PKcs) and a DNA-binding heterodimer (Ku70/80). To test whether DNA-PKcs activity is essential for CSR, we examined whether IgM+ B cells from scid mice with site-directed H and L chain transgenes were able to undergo CSR. Although B cells from these mice were shown to lack DNA-PKcs activity, they were able to switch from IgM to IgG or IgA with close to the same efficiency as B cells from control transgenic and nontransgenic scid/+ mice, heterozygous for the scid mutation. We conclude that CSR, unlike V(D)J recombination, can readily occur in the absence of DNA-PKcs activity. We suggest nonhomologous end joining may not be the (primary or only) mechanism used to repair DNA breaks during CSR.

scholarly journals DNA-PKcs phosphorylation at the T2609 cluster alters the repair pathway choice during immunoglobulin class switch recombination

2020 ◽  
Author(s):  
Jennifer L. Crowe ◽  
Xiaobin S. Wang ◽  
Zhengping Shao ◽  
Brian J. Lee ◽  
Verna Estes ◽  
...  
Keyword(s):  
Dna Repair ◽  
B Cells ◽  
Class Switch Recombination ◽  
Chromosomal Translocation ◽  
Radiation Sensitivity ◽  
Repair Pathway ◽  
End Joining ◽  
Class Switch ◽  
Pathway Choice

AbstractThe DNA-dependent protein kinase (DNA-PK), composed of the KU heterodimer and the large catalytic subunit (DNA-PKcs), is a classical non-homologous end-joining (cNHEJ) factor. Naïve B cells undergo class switch recombination (CSR) to generate antibodies with different isotypes by joining two DNA double-strand breaks at different switching regions via the cNHEJ pathway. DNA-PK and the cNHEJ pathway play important roles in the DNA repair phase of CSR. To initiate cNHEJ, KU binds to DNA ends, and recruits and activates DNA-PK. DNA-PKcs is the best-characterized substrate of DNA-PK, which phosphorylates DNA-PKcs at both the S2056 and T2609 clusters. Loss of T2609 cluster phosphorylation increases radiation sensitivity, suggesting a role of T2609 phosphorylation in DNA repair. Using the DNA-PKcs5A mouse model carrying an alanine substitution at the T2609 cluster, here we show that loss of T2609 phosphorylation of DNA-PKcs does not affect the CSR efficiency. Yet, the CSR junctions recovered from DNA-PKcs5A/5A B cells reveal increased chromosomal translocation, excess end-resection, and preferential usage of micro-homology – all signs of the alternative end-joining pathway. Thus, these results uncover a role of DNA-PKcs T2609 phosphorylation in promoting cNHEJ repair pathway choice during CSR.Key pointsLoss of T2069 cluster phosphorylation of DNA-PKcs promotes Alt-EJ-mediated CSR.

scholarly journals Histone methyltransferase MMSET promotes AID-mediated DNA breaks at the donor switch region during class switch recombination

2017 ◽  
Vol 114 (49) ◽  
pp. E10560-E10567 ◽  
Author(s):  
Hai Vu Nguyen ◽  
Junchao Dong ◽  
Rohit A. Panchakshari ◽  
Vipul Kumar ◽  
Frederick W. Alt ◽  
...  
Keyword(s):  
B Cells ◽  
Cytidine Deaminase ◽  
Class Switch Recombination ◽  
Histone Methyltransferase ◽  
Nonhomologous End Joining ◽  
Dna Double Strand Breaks ◽  
Dna Breaks ◽  
Set Domain ◽  
End Joining ◽  
Class Switch

In B cells, Ig class switch recombination (CSR) is initiated by activation-induced cytidine deaminase (AID), the activity of which leads to DNA double-strand breaks (DSBs) within IgH switch (S) regions. Preferential targeting of AID-mediated DSBs to S sequences is critical for allowing diversification of antibody functions, while minimizing potential off-target oncogenic events. Here, we used gene targeted inactivation of histone methyltransferase (HMT) multiple myeloma SET domain (MMSET) in mouse B cells and the CH12F3 cell line to explore its role in CSR. We find that deletion of MMSET-II, the isoform containing the catalytic SET domain, inhibits CSR without affecting either IgH germline transcription or joining of DSBs within S regions by classical nonhomologous end joining (C-NHEJ). Instead, we find that MMSET-II inactivation leads to decreased AID recruitment and DSBs at the upstream donor Sμ region. Our findings suggest a role for the HMT MMSET in promoting AID-mediated DNA breaks during CSR.

scholarly journals The Role of RNA in DNA Breaks, Repair and Chromosomal Rearrangements

Biomolecules ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 550
Author(s):  
Matvey Mikhailovich Murashko ◽  
Ekaterina Mikhailovna Stasevich ◽  
Anton Markovich Schwartz ◽  
Dmitriy Vladimirovich Kuprash ◽  
Aksinya Nicolaevna Uvarova ◽  
...  
Keyword(s):  
Chromosomal Rearrangements ◽  
Gene Mutations ◽  
Nonhomologous End Joining ◽  
Published Data ◽  
Dna Double Strand Breaks ◽  
Dna Breaks ◽  
End Joining ◽  
Homology Directed Repair ◽  
Chromosome Aberration Frequency

Incorrect reparation of DNA double-strand breaks (DSB) leading to chromosomal rearrangements is one of oncogenesis’s primary causes. Recently published data elucidate the key role of various types of RNA in DSB formation, recognition and repair. With growing interest in RNA biology, increasing RNAs are classified as crucial at the different stages of the main pathways of DSB repair in eukaryotic cells: nonhomologous end joining (NHEJ) and homology-directed repair (HDR). Gene mutations or variation in expression levels of such RNAs can lead to local DNA repair defects, increasing the chromosome aberration frequency. Moreover, it was demonstrated that some RNAs could stimulate long-range chromosomal rearrangements. In this review, we discuss recent evidence demonstrating the role of various RNAs in DSB formation and repair. We also consider how RNA may mediate certain chromosomal rearrangements in a sequence-specific manner.

scholarly journals Parp3 promotes long-range end joining in murine cells

2018 ◽  
Vol 115 (40) ◽  
pp. 10076-10081 ◽  
Author(s):  
Jacob V. Layer ◽  
J. Patrick Cleary ◽  
Alexander J. Brown ◽  
Kristen E. Stevenson ◽  
Sara N. Morrow ◽  
...  
Keyword(s):  
Chromosomal Rearrangements ◽  
Embryonic Stem ◽  
Cell Types ◽  
Strand Break ◽  
Nonhomologous End Joining ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Class Switch

Chromosomal rearrangements, including translocations, are early and essential events in the formation of many tumors. Previous studies that defined the genetic requirements for rearrangement formation have identified differences between murine and human cells, most notably in the role of classic and alternative nonhomologous end-joining (NHEJ) factors. We reported that poly(ADP)ribose polymerase 3 (PARP3) promotes chromosomal rearrangements induced by endonucleases in multiple human cell types. We show here that in contrast to classic (c-NHEJ) factors, Parp3 also promotes rearrangements in murine cells, including translocations in murine embryonic stem cells (mESCs), class–switch recombination in primary B cells, and inversions in tail fibroblasts that generateEml4–Alkfusions. In mESCs, Parp3-deficient cells had shorter deletion lengths at translocation junctions. This was corroborated using next-generation sequencing ofEml4–Alkjunctions in tail fibroblasts and is consistent with a role for Parp3 in promoting the processing of DNA double-strand breaks. We confirmed a previous report that Parp1 also promotes rearrangement formation. In contrast with Parp3, rearrangement junctions in the absence of Parp1 had longer deletion lengths, suggesting that Parp1 may suppress double-strand break processing. Together, these data indicate that Parp3 and Parp1 promote rearrangements with distinct phenotypes.

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