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| Status |
Public on Jul 16, 2015 |
| Title |
DNA double-strand breaks induced by AID in activated splenic B cells |
| Organism |
Mus musculus |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Activation-induced cytidine deaminase (AID) is required for initiation of Ig class switch recombination (CSR) and somatic hypermutation (SHM) of antibody genes during immune responses. AID has also been shown to induce chromosomal translocations, mutations, and DNA double-strand breaks (DSBs) involving non-Ig genes in activated B cells. To determine what makes a DNA site a target for AID-induced DSBs, we identify off-target DSBs induced by AID by performing chromatin immunoprecipitation (ChIP) for Nbs1, a protein that binds DSBs, followed by deep sequencing (ChIP-Seq). We detect and characterize hundreds of off-target AID-dependent DSBs. Two types of tandem repeats are highly enriched within the Nbs1-binding sites: long CA repeats, which can form Z-DNA, and tandem pentamers containing the AID target hotspot WGCW. These tandem repeats are not nearly as enriched at AID-independent DSBs, which we also identified. Msh2, a component of the mismatch repair pathway and important for genome stability, increases off-target DSBs, similar to its effect on Ig switch region DSBs, which are required intermediates during CSR. Most of the off-target DSBs are two-ended, consistent with generation during G1 phase, similar to DSBs in Ig switch regions. However, a minority are one-ended, presumably due to conversion of single-strand breaks to DSBs during replication. One-ended DSBs are repaired by processes involving homologous recombination, including break-induced replication repair, which can lead to genome instability. Off-target DSBs, especially those present during S phase, can lead to chromosomal translocations, deletions and gene amplifications, resulting in the high frequency of B cell lymphomas derived from cells that express or have expressed AID.
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| Overall design |
In two separate experiments, sites of Nbs1 binding were identified genomewide and compared in wild-type and AID-deficient splenic B cells undergoing class switch recombination.
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| Contributor(s) |
Khair L, Baker RE, Linehan EK, Schrader CE, Stavnezer J |
| Citation(s) |
26263206 |
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| Submission date |
Mar 02, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Richard E Baker |
| E-mail(s) |
richard.baker@umassmed.edu
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| Phone |
508-856-6046
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| Organization name |
University of Massachusetts Medical School
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| Department |
Microbiology & Physiological Systems
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| Street address |
55 Lake Avenue North
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| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01655 |
| Country |
USA |
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| Platforms (2) |
| GPL11002 |
Illumina Genome Analyzer IIx (Mus musculus) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
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| Samples (7)
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| Relations |
| BioProject |
PRJNA276893 |
| SRA |
SRP055714 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE66424_RAW.tar |
1.7 Gb |
(http)(custom) |
TAR (of BED, BEDGRAPH) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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