The goal of this research project is to further characterize the mechanisms and regulation of antibody gene isotype switching. Three quite distinct mechanisms have been postulated to be important in isotype switching: 1) DNA switch recombination; 2) alternative RNA processing of long transcripts; and 3) RNA trans-splicing. Strong evidence supports intrachromosomal DNA recombination/deletion as an important mechanism involved in antibody gene class switching. However, we have found that a murine antibody mu heavy chain transgene can undergo isotype switching and produce large amounts of IgG in response to immunization with the appropriate antigen. Surprisingly, this transgene isotype switching reflects interchromosomal DNA recombination events between the transgene and the endogenous heavy chain gene locus which result in chromosomal translocations and which appear to be mediated by the normal class switch recombination mechanisms. Other laboratories analyzing isotype switching of a human antibody heavy chain transgene have reported that transgene switching can apparently occur by an RNA trans-splicing mechanism. We will further characterize transgene switching in lymphomas hybridomas, and B-cells derived from our transgenic mice to explore the interchromosomal mechanisms responsible for transgene switching. We will also investigate interchromosomal isotype switching of antibody heavy chain genes in non-transgenic mice. A component of these studies will involve investigating the relative frequencies of intrachromosomal and interchromosomal switching. In addition, we will use deletional mutagenesis to characterize DNA sequence elements important for the isotype switching process both in transgenic mice and in mutant mice derived by homologous recombination mutagenesis in embryonic stem (ES) cells.
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