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Heinz Jacobs - One of the best experts on this subject based on the ideXlab platform.
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a t mutaGenesis in hypermutated Immunoglobulin Genes strongly depends on pcnak164 modification
Journal of Experimental Medicine, 2007Co-Authors: Petra Langerak, Anders O H Nygren, Peter H L Krijger, Paul C M Van Den Berk, Heinz JacobsAbstract:B cells use translesion DNA synthesis (TLS) to introduce somatic mutations around genetic lesions caused by activation-induced cytidine deaminase. Monoubiquitination at lysine164 of proliferating cell nuclear antigen (PCNAK164) stimulates TLS. To determine the role of PCNAK164 modifications in somatic hypermutation, PCNAK164R knock-in mice were generated. PCNAK164R/K164R mutants are born at a sub-Mendelian frequency. Although PCNAK164R/K164R B cells proliferate and class switch normally, the mutation spectrum of hypermutated Immunoglobulin (Ig) Genes alters dramatically. A strong reduction of mutations at template A/T is associated with a compensatory increase at G/C, which is a phenotype similar to polymerase η (Polη) and mismatch repair–deficient B cells. Mismatch recognition, monoubiquitinated PCNA, and Polη likely cooperate in establishing mutations at template A/T during replication of Ig Genes.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Paul C M Van Den Berk, Heinz Jacobs, Niels De WindAbstract:DNA translesion synthesis (TLS) is a backup replication pathway that, in contrast to replicative polymerases δ and e, is capable of replicating damaged nucleotides that confer helical distortion to the DNA template. Replication of the damaged nucleotide by TLS is believed to safeguard the perpetuation of replication in the presence of unrepaired DNA damage, albeit frequently at the expense of misincorporations. The Y family of DNA polymerases in mammals is a major class of TLS polymerases comprising the polymerases η, ι, κ, and Rev1 (1). In vitro, the catalytic activity of mammalian Rev1 is limited to the highly distributive incorporation of cytosine residues opposite deoxyuridine residues and abasic nucleotides (2, 3). Analysis of TLS at site-specifically damaged DNA templates in Saccharomyces cerevisiae supports an important role of Rev1 in the bypass of abasic sites in vivo (4). In addition, Rev1-deficient chicken DT40 cells and hypomorphic mouse Rev1 mutant cells display hypersensitivity to a variety of genotoxic agents (5, 6). The infrequent mutations to deoxycytidine induced by these agents in Rev1-proficient S. cerevisiae suggests a second (noncatalytic) role for Rev1, possibly by recruiting other TLS polymerases. In agreement, TLS polymerases η, ι, κ, as well as the Rev7 TLS-associated protein, interact with a COOH-terminal domain of Rev1 (references 7–10; unpublished data). Deoxyuridine and abasic sites are essential triggers for somatic hypermutation (SHM), a process of antibody diversification in which the variable regions of Ig heavy (IgH) and light (IgL) chain Genes in proliferating B cells of the germinal center mutate at an extremely high rate (11). This is followed by clonal selection of the cells that express Ig with increased affinity toward the antigen (12). SHM is triggered by deamination to uracil of deoxycytidines within Ig Genes by the activation-induced deoxycytidine deaminase (AID) (11, 13). Subsequent processing by uracil DNA glycosylase (UNG) can generate abasic sites that may be bypassed by one or more of the TLS polymerases (14). In a second phase of SHM, DNA mismatch repair may induce single-stranded gaps at sites of mispaired deoxyuridine residues, followed by filling the gaps by mutagenic TLS (11, 15). To investigate involvement of the Rev1 TLS polymerase in SHM, we have generated and analyzed Rev1-deficient mice.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Heinz Jacobs, Paul C M Van Den Berk, Niels De WindAbstract:Somatic hypermutation of Ig Genes enables B cells of the germinal center to generate high-affinity Immunoglobulin variants. Key intermediates in somatic hypermutation are deoxyuridine lesions, introduced by activation-induced cytidine deaminase. These lesions can be processed further to abasic sites by uracil DNA glycosylase. Mutagenic replication of deoxyuridine, or of its abasic derivative, by translesion synthesis polymerases is hypothesized to underlie somatic hypermutation. Rev1 is a translesion synthesis polymerase that in vitro incorporates uniquely deoxycytidine opposite deoxyuridine and abasic residues. To investigate a role of Rev1 in mammalian somatic hypermutation we have generated mice deficient for Rev1. Although Rev1-/- mice display transient growth retardation, proliferation of Rev1-/- LPS-stimulated B cells is indistinguishable from wild-type cells. In mutated Ig Genes from Rev1-/- mice, C to G transversions were virtually absent in the nontranscribed (coding) strand and reduced in the transcribed strand. This defect is associated with an increase of A to T, C to A, and T to C substitutions. These results indicate that Rev1 incorporates deoxycytidine residues, most likely opposite abasic nucleotides, during somatic hypermutation. In addition, loss of Rev1 causes compensatory increase in mutaGenesis by other translesion synthesis polymerases.
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dna double strand breaks in Immunoglobulin Genes undergoing somatic hypermutation
Immunity, 2000Co-Authors: Linda Bross, Yosho Fukita, Fraser Mcblane, Corinne Demolliere, Klaus Rajewsky, Heinz JacobsAbstract:How rearranged Immunoglobulin (Ig) Genes are further diversified by somatic hypermutation is unknown. Using VDJ passenger Ig heavy chain (IgH) knockin mouse strains, we now demonstrate a high frequency of DNA double-strand breaks (DSBs) in the targeted VDJ passenger gene of germinal center (GC) B cells. These DSBs parallel the distribution of mutations in the targeted hypermutation domain and are found preferentially at RGYW motifs, the intrinsic hot spots of somatic hypermutation. The introduction of DSBs appears to depend on transcriptional activity. Thus, secondary diversification of rearranged V gene segments relates to an error-prone nonhomologous DSB repair system acting in B cells of the GC.
Niels De Wind - One of the best experts on this subject based on the ideXlab platform.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Paul C M Van Den Berk, Heinz Jacobs, Niels De WindAbstract:DNA translesion synthesis (TLS) is a backup replication pathway that, in contrast to replicative polymerases δ and e, is capable of replicating damaged nucleotides that confer helical distortion to the DNA template. Replication of the damaged nucleotide by TLS is believed to safeguard the perpetuation of replication in the presence of unrepaired DNA damage, albeit frequently at the expense of misincorporations. The Y family of DNA polymerases in mammals is a major class of TLS polymerases comprising the polymerases η, ι, κ, and Rev1 (1). In vitro, the catalytic activity of mammalian Rev1 is limited to the highly distributive incorporation of cytosine residues opposite deoxyuridine residues and abasic nucleotides (2, 3). Analysis of TLS at site-specifically damaged DNA templates in Saccharomyces cerevisiae supports an important role of Rev1 in the bypass of abasic sites in vivo (4). In addition, Rev1-deficient chicken DT40 cells and hypomorphic mouse Rev1 mutant cells display hypersensitivity to a variety of genotoxic agents (5, 6). The infrequent mutations to deoxycytidine induced by these agents in Rev1-proficient S. cerevisiae suggests a second (noncatalytic) role for Rev1, possibly by recruiting other TLS polymerases. In agreement, TLS polymerases η, ι, κ, as well as the Rev7 TLS-associated protein, interact with a COOH-terminal domain of Rev1 (references 7–10; unpublished data). Deoxyuridine and abasic sites are essential triggers for somatic hypermutation (SHM), a process of antibody diversification in which the variable regions of Ig heavy (IgH) and light (IgL) chain Genes in proliferating B cells of the germinal center mutate at an extremely high rate (11). This is followed by clonal selection of the cells that express Ig with increased affinity toward the antigen (12). SHM is triggered by deamination to uracil of deoxycytidines within Ig Genes by the activation-induced deoxycytidine deaminase (AID) (11, 13). Subsequent processing by uracil DNA glycosylase (UNG) can generate abasic sites that may be bypassed by one or more of the TLS polymerases (14). In a second phase of SHM, DNA mismatch repair may induce single-stranded gaps at sites of mispaired deoxyuridine residues, followed by filling the gaps by mutagenic TLS (11, 15). To investigate involvement of the Rev1 TLS polymerase in SHM, we have generated and analyzed Rev1-deficient mice.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Heinz Jacobs, Paul C M Van Den Berk, Niels De WindAbstract:Somatic hypermutation of Ig Genes enables B cells of the germinal center to generate high-affinity Immunoglobulin variants. Key intermediates in somatic hypermutation are deoxyuridine lesions, introduced by activation-induced cytidine deaminase. These lesions can be processed further to abasic sites by uracil DNA glycosylase. Mutagenic replication of deoxyuridine, or of its abasic derivative, by translesion synthesis polymerases is hypothesized to underlie somatic hypermutation. Rev1 is a translesion synthesis polymerase that in vitro incorporates uniquely deoxycytidine opposite deoxyuridine and abasic residues. To investigate a role of Rev1 in mammalian somatic hypermutation we have generated mice deficient for Rev1. Although Rev1-/- mice display transient growth retardation, proliferation of Rev1-/- LPS-stimulated B cells is indistinguishable from wild-type cells. In mutated Ig Genes from Rev1-/- mice, C to G transversions were virtually absent in the nontranscribed (coding) strand and reduced in the transcribed strand. This defect is associated with an increase of A to T, C to A, and T to C substitutions. These results indicate that Rev1 incorporates deoxycytidine residues, most likely opposite abasic nucleotides, during somatic hypermutation. In addition, loss of Rev1 causes compensatory increase in mutaGenesis by other translesion synthesis polymerases.
David G Schatz - One of the best experts on this subject based on the ideXlab platform.
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uracil residues dependent on the deaminase aid in Immunoglobulin gene variable and switch regions
Nature Immunology, 2011Co-Authors: Robert W Maul, David G Schatz, Stella A Martomo, Huseyin Saribasak, Rhonda L Mcclure, William Yang, Alexandra Vaisman, Hillary S Gramlich, Roger Woodgate, David M WilsonAbstract: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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somatic hypermutation of Immunoglobulin Genes merging mechanisms for genetic diversity
Cell, 2002Co-Authors: Nina F Papavasiliou, David G SchatzAbstract:Somatic hypermutation is critical for the generation of high-affinity antibodies and effective immune responses, but its molecular mechanism remains poorly understood. Recent studies have identified DNA strand lesions associated with the hypermutation process and suggested the involvement of specific repair molecules and pathways. Particularly exciting has been the discovery of a putative RNA editing enzyme, the activation-induced cytidine deaminase (AID), that is required for all Immunoglobulin gene-specific modification reactions (somatic hypermutation, class switch recombination, and gene conversion). Parallels between these three reactions are considered in light of recent advances.
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cell cycle regulated dna double strand breaks in somatic hypermutation of Immunoglobulin Genes
Nature, 2000Co-Authors: Nina F Papavasiliou, David G SchatzAbstract:Cell-cycle-regulated DNA double-strand breaks in somatic hypermutation of Immunoglobulin Genes
Jacob G Jansen - One of the best experts on this subject based on the ideXlab platform.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Paul C M Van Den Berk, Heinz Jacobs, Niels De WindAbstract:DNA translesion synthesis (TLS) is a backup replication pathway that, in contrast to replicative polymerases δ and e, is capable of replicating damaged nucleotides that confer helical distortion to the DNA template. Replication of the damaged nucleotide by TLS is believed to safeguard the perpetuation of replication in the presence of unrepaired DNA damage, albeit frequently at the expense of misincorporations. The Y family of DNA polymerases in mammals is a major class of TLS polymerases comprising the polymerases η, ι, κ, and Rev1 (1). In vitro, the catalytic activity of mammalian Rev1 is limited to the highly distributive incorporation of cytosine residues opposite deoxyuridine residues and abasic nucleotides (2, 3). Analysis of TLS at site-specifically damaged DNA templates in Saccharomyces cerevisiae supports an important role of Rev1 in the bypass of abasic sites in vivo (4). In addition, Rev1-deficient chicken DT40 cells and hypomorphic mouse Rev1 mutant cells display hypersensitivity to a variety of genotoxic agents (5, 6). The infrequent mutations to deoxycytidine induced by these agents in Rev1-proficient S. cerevisiae suggests a second (noncatalytic) role for Rev1, possibly by recruiting other TLS polymerases. In agreement, TLS polymerases η, ι, κ, as well as the Rev7 TLS-associated protein, interact with a COOH-terminal domain of Rev1 (references 7–10; unpublished data). Deoxyuridine and abasic sites are essential triggers for somatic hypermutation (SHM), a process of antibody diversification in which the variable regions of Ig heavy (IgH) and light (IgL) chain Genes in proliferating B cells of the germinal center mutate at an extremely high rate (11). This is followed by clonal selection of the cells that express Ig with increased affinity toward the antigen (12). SHM is triggered by deamination to uracil of deoxycytidines within Ig Genes by the activation-induced deoxycytidine deaminase (AID) (11, 13). Subsequent processing by uracil DNA glycosylase (UNG) can generate abasic sites that may be bypassed by one or more of the TLS polymerases (14). In a second phase of SHM, DNA mismatch repair may induce single-stranded gaps at sites of mispaired deoxyuridine residues, followed by filling the gaps by mutagenic TLS (11, 15). To investigate involvement of the Rev1 TLS polymerase in SHM, we have generated and analyzed Rev1-deficient mice.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Heinz Jacobs, Paul C M Van Den Berk, Niels De WindAbstract:Somatic hypermutation of Ig Genes enables B cells of the germinal center to generate high-affinity Immunoglobulin variants. Key intermediates in somatic hypermutation are deoxyuridine lesions, introduced by activation-induced cytidine deaminase. These lesions can be processed further to abasic sites by uracil DNA glycosylase. Mutagenic replication of deoxyuridine, or of its abasic derivative, by translesion synthesis polymerases is hypothesized to underlie somatic hypermutation. Rev1 is a translesion synthesis polymerase that in vitro incorporates uniquely deoxycytidine opposite deoxyuridine and abasic residues. To investigate a role of Rev1 in mammalian somatic hypermutation we have generated mice deficient for Rev1. Although Rev1-/- mice display transient growth retardation, proliferation of Rev1-/- LPS-stimulated B cells is indistinguishable from wild-type cells. In mutated Ig Genes from Rev1-/- mice, C to G transversions were virtually absent in the nontranscribed (coding) strand and reduced in the transcribed strand. This defect is associated with an increase of A to T, C to A, and T to C substitutions. These results indicate that Rev1 incorporates deoxycytidine residues, most likely opposite abasic nucleotides, during somatic hypermutation. In addition, loss of Rev1 causes compensatory increase in mutaGenesis by other translesion synthesis polymerases.
Petra Langerak - One of the best experts on this subject based on the ideXlab platform.
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a t mutaGenesis in hypermutated Immunoglobulin Genes strongly depends on pcnak164 modification
Journal of Experimental Medicine, 2007Co-Authors: Petra Langerak, Anders O H Nygren, Peter H L Krijger, Paul C M Van Den Berk, Heinz JacobsAbstract:B cells use translesion DNA synthesis (TLS) to introduce somatic mutations around genetic lesions caused by activation-induced cytidine deaminase. Monoubiquitination at lysine164 of proliferating cell nuclear antigen (PCNAK164) stimulates TLS. To determine the role of PCNAK164 modifications in somatic hypermutation, PCNAK164R knock-in mice were generated. PCNAK164R/K164R mutants are born at a sub-Mendelian frequency. Although PCNAK164R/K164R B cells proliferate and class switch normally, the mutation spectrum of hypermutated Immunoglobulin (Ig) Genes alters dramatically. A strong reduction of mutations at template A/T is associated with a compensatory increase at G/C, which is a phenotype similar to polymerase η (Polη) and mismatch repair–deficient B cells. Mismatch recognition, monoubiquitinated PCNA, and Polη likely cooperate in establishing mutations at template A/T during replication of Ig Genes.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Paul C M Van Den Berk, Heinz Jacobs, Niels De WindAbstract:DNA translesion synthesis (TLS) is a backup replication pathway that, in contrast to replicative polymerases δ and e, is capable of replicating damaged nucleotides that confer helical distortion to the DNA template. Replication of the damaged nucleotide by TLS is believed to safeguard the perpetuation of replication in the presence of unrepaired DNA damage, albeit frequently at the expense of misincorporations. The Y family of DNA polymerases in mammals is a major class of TLS polymerases comprising the polymerases η, ι, κ, and Rev1 (1). In vitro, the catalytic activity of mammalian Rev1 is limited to the highly distributive incorporation of cytosine residues opposite deoxyuridine residues and abasic nucleotides (2, 3). Analysis of TLS at site-specifically damaged DNA templates in Saccharomyces cerevisiae supports an important role of Rev1 in the bypass of abasic sites in vivo (4). In addition, Rev1-deficient chicken DT40 cells and hypomorphic mouse Rev1 mutant cells display hypersensitivity to a variety of genotoxic agents (5, 6). The infrequent mutations to deoxycytidine induced by these agents in Rev1-proficient S. cerevisiae suggests a second (noncatalytic) role for Rev1, possibly by recruiting other TLS polymerases. In agreement, TLS polymerases η, ι, κ, as well as the Rev7 TLS-associated protein, interact with a COOH-terminal domain of Rev1 (references 7–10; unpublished data). Deoxyuridine and abasic sites are essential triggers for somatic hypermutation (SHM), a process of antibody diversification in which the variable regions of Ig heavy (IgH) and light (IgL) chain Genes in proliferating B cells of the germinal center mutate at an extremely high rate (11). This is followed by clonal selection of the cells that express Ig with increased affinity toward the antigen (12). SHM is triggered by deamination to uracil of deoxycytidines within Ig Genes by the activation-induced deoxycytidine deaminase (AID) (11, 13). Subsequent processing by uracil DNA glycosylase (UNG) can generate abasic sites that may be bypassed by one or more of the TLS polymerases (14). In a second phase of SHM, DNA mismatch repair may induce single-stranded gaps at sites of mispaired deoxyuridine residues, followed by filling the gaps by mutagenic TLS (11, 15). To investigate involvement of the Rev1 TLS polymerase in SHM, we have generated and analyzed Rev1-deficient mice.
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strand biased defect in c g transversions in hypermutating Immunoglobulin Genes in rev1 deficient mice
Journal of Experimental Medicine, 2006Co-Authors: Jacob G Jansen, Anastasia Tsaalbishtylik, Petra Langerak, Heinz Jacobs, Paul C M Van Den Berk, Niels De WindAbstract:Somatic hypermutation of Ig Genes enables B cells of the germinal center to generate high-affinity Immunoglobulin variants. Key intermediates in somatic hypermutation are deoxyuridine lesions, introduced by activation-induced cytidine deaminase. These lesions can be processed further to abasic sites by uracil DNA glycosylase. Mutagenic replication of deoxyuridine, or of its abasic derivative, by translesion synthesis polymerases is hypothesized to underlie somatic hypermutation. Rev1 is a translesion synthesis polymerase that in vitro incorporates uniquely deoxycytidine opposite deoxyuridine and abasic residues. To investigate a role of Rev1 in mammalian somatic hypermutation we have generated mice deficient for Rev1. Although Rev1-/- mice display transient growth retardation, proliferation of Rev1-/- LPS-stimulated B cells is indistinguishable from wild-type cells. In mutated Ig Genes from Rev1-/- mice, C to G transversions were virtually absent in the nontranscribed (coding) strand and reduced in the transcribed strand. This defect is associated with an increase of A to T, C to A, and T to C substitutions. These results indicate that Rev1 incorporates deoxycytidine residues, most likely opposite abasic nucleotides, during somatic hypermutation. In addition, loss of Rev1 causes compensatory increase in mutaGenesis by other translesion synthesis polymerases.