Deoxyuridine Triphosphate Incorporation during Somatic Hypermutation of Mouse <i>VkOx</i> Genes after Immunization with Phenyloxazolone

Roche, Benjamin; Claës, Aurélie; Rougeon, François

doi:10.4049/jimmunol.1001459

Cited by 8 publications

(11 citation statements)

References 33 publications

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“…It has been proposed that a DNA polymerase might use this non-canonical base from the nucleotide pool to fill gaps produced during SHM 39,40 . To test that hypothesis, we first did DNA-replication assays on a primer-template substrate (Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

Uracil residues dependent on the deaminase AID in immunoglobulin gene variable and switch regions

et al. 2010

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Activation-induced deaminase (AID) initiates diversity of immunoglobulin genes through deamination of cytosine to uracil. Two opposing models have been proposed for the deamination of DNA or RNA by AID. Although most data support DNA deamination, there is no physical evidence of uracil residues in immunoglobulin genes. Here we demonstrate their presence by determining the sensitivity of DNA to digestion with uracil DNA glycosylase (UNG) and abasic endonuclease. Using several methods of detection, we identified uracil residues in the variable and switch regions. Uracil residues were generated within 24 h of B cell stimulation, were present on both DNA strands and were found to replace mainly cytosine bases. Our data provide direct evidence for the model that AID functions by deaminating cytosine residues in DNA.

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Section: Resultsmentioning

confidence: 99%

Uracil residues dependent on the deaminase AID in immunoglobulin gene variable and switch regions

et al. 2010

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“…Although directed cytosine deamination by the enzyme AID is the major source of uracil during these processes, direct incorporation of dUTP by DNA polymerase has not yet been ruled out as a minor contributor 107 . It was recently reported, for example, that uracil can be detected at positions of thymines in the variable region of mouse immunoglobulin genes 108 . Another study concluded, however, that uracil in hypermutating immunoglobulin genes was only found at positions of cytosines 109 .…”

Section: Transcription-associated Changes In Dna Compositionmentioning

confidence: 99%

Transcription as a source of genome instability

2012

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Alterations in genome sequence and structure contribute to somatic disease, affect the fitness of subsequent generations and drive evolutionary processes. The critical roles of highly accurate replication and efficient repair in maintaining overall genome integrity are well known, but the more localized stability costs associated with transcribing DNA into RNA molecules are less appreciated. Here we review the diverse ways that the essential process of transcription alters the underlying DNA template and thereby modifies the genetic landscape.

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“…The only reports of uracils created by AID in normal B cells have been in mouse models of antibody maturation that have focused only on the Ig genes (25,26). Furthermore, the balance between uracil creation in the B cell genome by AID and its removal by UNG2 or other repair enzymes has not been examined.…”

mentioning

confidence: 99%

Genomic Uracil Homeostasis during Normal B Cell Maturation and Loss of This Balance during B Cell Cancer Development

Shalhout

Haddad

Sosin

et al. 2014

Molecular and Cellular Biology

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d Activation-induced deaminase (AID) converts DNA cytosines to uracils in immunoglobulin genes, creating antibody diversification. It also causes mutations and translocations that promote cancer. We examined the interplay between uracil creation by AID and its removal by UNG2 glycosylase in splenocytes undergoing maturation and in B cell cancers. The genomic uracil levels remain unchanged in normal stimulated B cells, demonstrating a balance between uracil generation and removal. In stimulated UNG ؊/؊ cells, uracil levels increase by 11-to 60-fold during the first 3 days. In wild-type B cells, UNG2 gene expression and enzymatic activity rise and fall with AID levels, suggesting that UNG2 expression is coordinated with uracil creation by AID. Remarkably, a murine lymphoma cell line, several human B cell cancer lines, and human B cell tumors expressing AID at high levels have genomic uracils comparable to those seen with stimulated UNG ؊/؊ splenocytes. However, cancer cells express UNG2 gene at levels similar to or higher than those seen with peripheral B cells and have nuclear uracil excision activity comparable to that seen with stimulated wild-type B cells. We propose that more uracils are created during B cell cancer development than are removed from the genome but that the uracil creation/excision balance is restored during establishment of cell lines, fixing the genomic uracil load at high levels. When B lymphocytes are activated through antigen presentation, they acquire hypermutations in the immunoglobulin (Ig) genes, facilitating affinity maturation of antibodies. An enzyme, activation-induced deaminase (AID), initiates these events by converting cytosines in DNA to uracil (1-4). The introduction of this rare base into DNA leads to a very high frequency of base substitution mutations in the Ig variable domain (known as somatic hypermutations [SHMs]; reviewed in references 5 and 6). Generation of uracils is also the first step in the creation of strand breaks in the switch regions of Ig genes, leading to the replacement of the constant domain with other domains such as ␥, in a process called class-switch recombination (CSR; reviewed in reference 7). AID also binds near the transcription start sites of a large number of actively transcribed genes (8) and mutates a number of additional genes, including those encoding BCL-6, MYC, PAX-5, and PIM1 (9-12). The uracils generated by AID are thought to be removed by the nuclear form of the uracil-DNA glycosylase, UNG2, creating abasic sites that are repaired by error-prone copying mechanisms causing hypermutations (13,14). Another uracil-DNA glycosylase, SMUG1, is normally considered the backup enzyme for UNG2 (15), but overproduction of SMUG1 is required for it to complement an UNG Ϫ/Ϫ mutant during CSR (16). Additionally, DNA mismatch repair (MMR) also plays a role in shaping the mutation spectrum in SHM (17).There is a strong connection between expression of AID and cancers in animals. Constitutive expression of AID in mice causes T cell cancers (18). Many human...

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Deoxyuridine Triphosphate Incorporation during Somatic Hypermutation of Mouse VkOx Genes after Immunization with Phenyloxazolone

Cited by 8 publications

References 33 publications

Uracil residues dependent on the deaminase AID in immunoglobulin gene variable and switch regions

Uracil residues dependent on the deaminase AID in immunoglobulin gene variable and switch regions

Transcription as a source of genome instability

Genomic Uracil Homeostasis during Normal B Cell Maturation and Loss of This Balance during B Cell Cancer Development

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