Msh2 ATPase Activity Is Essential for Somatic Hypermutation at A-T Basepairs and for Efficient Class Switch Recombination

Martín, Alberto; Li, Ziqiang; Lin, Diana P.; Bardwell, Philip D.; Iglesias-Ussel, Maria D.; Edelmann, Winfried; Scharff, Matthew D.

doi:10.1084/jem.20030880

Cited by 95 publications

(97 citation statements)

References 50 publications

(99 reference statements)

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“…Most of the mutations were transitions and in hotspots and all were in G:C base pairs (Table 1). In most cultured B cells approximately 80% of the mutations are in G:C base pairs (Martin et al, 2002a), which is higher than is seen in vivo in humans or mice (Martin et al, 2003;Li et al, 2004). However, it is unusual to see no mutations in A:T base pairs (Green et al, 1998), although we have seen this in another hybridoma (Martin et al, 2002a).…”

Section: Aid Induces V Region Mutations In Hybridomas and Antigen Binmentioning

confidence: 45%

Forced expression of AID facilitates the isolation of class switch variants from hybridoma cells

Iglesias-Ussel

Fan

et al. 2006

Journal of Immunological Methods

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Monoclonal antibodies are used in the treatment and diagnosis of diseases and to study the protective and adverse functions of antibodies in vitro and in vivo. Since the isotype determines the effector function, half-life in the serum and distribution throughout the body, it would be useful to have a battery of antibodies with the same binding site associated with different isotypes. However, since hybridomas switch isotypes at very low frequencies in tissue culture, it has been difficult and very labor intensive to isolate panels of class switch variants. We show here that stable transfection of activation-induced cytidine deaminase (AID) in hybridomas increased their frequency of switching to a level that greatly facilitated the isolation of subclones expressing monoclonal antibodies of different isotypes. Although forced expression of AID also increased the frequency of somatic hypermutation in the immunoglobulin variable regions that encode the antigen-binding site, antigen recognition was retained in the isotype switched antibodies.

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Section: Aid Induces V Region Mutations In Hybridomas and Antigen Binmentioning

confidence: 45%

Forced expression of AID facilitates the isolation of class switch variants from hybridoma cells

Iglesias-Ussel

Fan

et al. 2006

Journal of Immunological Methods

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“…The fact that NHEJ is important for CSR but not for SHM supports the idea that AID-initiated G-U mismatches are resolved differently during SHM and CSR (for review, see Li et al 2003). Although H2AX-deficient mice exhibit normal SHM (Reina-San- Martin et al 2003), in hypermutating Burkitt's lymphoma cell lines ␥-H2AX is associated with the V region but not C region Zan et al 2003). This suggests that double-stranded breaks may arise during SHM but are not required for that process (Faili et al 2002b;Papavasiliou and Schatz 2002), whereas they are essential for repairing the double-stranded DNA breaks that are a critical part of CSR.…”

Section: Nonhomologous End Joiningmentioning

confidence: 70%

“…2D), allowing each antigen binding site to mediate many different effector functions. It is important to note that point mutations with the similar characteristics to those seen in V region SHM are found in the donor and recipient S regions flanking the site of recombination (ReinaSan- Martin et al 2003).…”

Section: Characteristics Of Class Switch Recombinationmentioning

confidence: 99%

The generation of antibody diversity through somatic hypermutation and class switch recombination

et al. 2004

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“…RNAP II was immunoprecipitated with an antibody that does not discriminate between different modifications of the C-terminal domain of RNAP II, so the ChIP results would reflect the number of RNAP II molecules associated with the examined DNA sequence (34). The immunoprecipitated DNA was amplified with primers specific for each of the S regions that generated Ϸ100-bp PCR fragments (see Materials and Methods) from regions that undergo high rates of recombination (33,(35)(36)(37)(38)(39). At each time point and each treatment, the ChIP was done with similar amounts of chromatin as demonstrated by the PCR products of the DNA before immunoprecipitation (Fig.…”

Section: Resultsmentioning

confidence: 99%

Differential regulation of histone acetylation and generation of mutations in switch regions is associated with Ig class switching

Luo

Scharff

2004

Proc. Natl. Acad. Sci. U.S.A.

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Class switch recombination (CSR) allows B cells to make effective protective antibodies. CSR involves the replacement of the constant region with one of the downstream constant regions by recombination between the donor and recipient switch (S) regions. Although histone H3 hyperacetylation in recipient S regions was recently reported to coincide with CSR, the relative histone H3 and H4 acetylation status of the donor and recipient S regions and the relationship between the generation of mutations and histone hyperacetylation in S regions have not been addressed. Here we report that histone H3 and H4 were constitutively hyperacetylated in the donor S region before and after different mitogen and cytokine treatments. We observed an increased frequency of mutations in hyperacetylated S␥ DNA segments immunoprecipitated with anti-acetyl histone antibodies. Furthermore, time course experiments revealed that the pattern of association of RNA polymerase II with S regions was much like that of H3 hyperacetylation but not always like that of H4 hyperacetylation. Collectively, our data suggest that H3 and H4 histone hyperacetylation in different S regions is regulated differently, that RNA polymerase II distribution and H3 hyperacetylation reflect the transcriptional activity of a given S region, and that transcription, hyperacetylation, and mutation are not sufficient to guarantee CSR. These findings support the notion that there are additional modifications and͞or factors involved in the complex process of CSR.H igh-affinity IgG, IgA, and IgE antibodies protect higher organisms from infection by pathogenic organisms and other environmental threats. The generation of effective protective antibodies requires B cells to carry out somatic hypermutation (SHM) and class switch recombination (CSR), two related but quite different DNA transactions (1). SHM introduces many point mutations in the variable (V) regions of the Ig heavy and light chain genes that encode the antigen-binding site of the antibody molecule (2). In contrast to SHM, CSR is a region-specific recombinationdeletion process that requires the generation of double-stranded DNA breaks (DSB) (3). These DSB are generated in the donor switch (S) region that is just upstream of the constant (C) region gene and in a downstream recipient ␥, , or ␣ S region (4, 5). Recombination then brings one of those downstream C regions into proximity to the V region. This allows the mutated heavy chain V region to be expressed with one of the C regions so that each antigen-binding site will be able to mediate different effector functions and be distributed throughout the body.Despite the different outcomes of SHM and CSR, activationinduced cytidine deaminase (AID) is required for both processes (6) presumably because of its ability to deaminate deoxycytidine to deoxyuridine on single-stranded DNA (ssDNA) (7-11). Both SHM and CSR require transcription (12), suggesting that the ssDNA substrate for AID is created by the generation of transcription bubbles (7) and͞or perhaps triplex RNA...

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Msh2 ATPase Activity Is Essential for Somatic Hypermutation at A-T Basepairs and for Efficient Class Switch Recombination

Cited by 95 publications

References 50 publications

Forced expression of AID facilitates the isolation of class switch variants from hybridoma cells

Forced expression of AID facilitates the isolation of class switch variants from hybridoma cells

The generation of antibody diversity through somatic hypermutation and class switch recombination

Differential regulation of histone acetylation and generation of mutations in switch regions is associated with Ig class switching

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