The aim of this research program is to characterize immunoglobulin switch recombination in normal B lymphocytes. Mitogen stimulation of spleen cell cultures induces intra-switch (S mu) region rearrangements which effect up to 80% of S mu DNA. The high degree and inducibility of intra-S DNA recombination permits the molecular and biochemical analysis of this recombination event. The generality of intra-S DNA recombination in switch recombination will be tested using various inducers of B cell activation. The regulation isotype expression will be examined by correlating the degree and timing of intra-S mu and intra-S delta recombination with administration of specific B cell growth factors known to influence the spectrum of isotype response. The structure and DNA sequence of recombinant intra-S DNA molecules will be studied. This information may focus attention on hitherto unrecognized secondary structure which is involved in this event. Chromatin structure, as measured by DNasel hypersensitive sites, methylation status and nucleosome phasing, will be examined to determine its effect on switch recombination in normal B cells. The dependency of intra-S DNA recombination on transcriptional competence will be investigated by construction of a transgenic mouse with marked immunoglobulin genes under inducible control. Proteins which specifically bind S DNA and induced during intra-S DNA recombination will be identified and characterized for their role in switch recombination regulation.
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