Class switch DNA recombinations are important for isotype switching in B-cells and also appear to be involved in chromosomal translocations of some oncogenes However, little is known bout the switching mechanism. Tandemly-repeated sequences located upstream of all H-chain antibody constant region genes have generally been thought to be important in the targeting and/or the mechanism of class switch recombination. We have used gene targeting to delete the tandem repeats from the switch (S) region associated with the C-mu constant region in mice. Surprisingly, these mutant mice are still able to undergo isotype switching, though the efficiency of the process is reduced. We also find that switching to different isotypes is differentially affected by deletion of the S-mu tandem repeats. The current application seeks to extend our preliminary results to analyze several aspects of class switch DNA recombination. First, the mice lacking S-mu will be analyzed to determine whether IgA-switching is essentially intact as indicated by preliminary data and to assess the effect of S-mu deletion of IgE switching. Potential molecular explanations for the different effects of S-mu on switching to the different isotypes will be investigated by characterizing switch recombination junction sites for isotypes that show the greatest and least effect due to S-mu deletion. Second , double stranded DNA breaks within the JH-C mu intron will be analyzed in wild-type and S-mu mutant mice to assess whether DNA cleavage sites might be affected by S-mu deletion. Third, gene targeting will be used to delete additional sequences within the JH-C mu intron to localize those sequences that are required for isotype switching. And fourth, specific DNA sequences will be introduce by gene targeting into an IgH allele that is defective for switching to investigate the minimal sequences that can restore the capacity for switching.
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