Secondary deletional recombination of rearranged switch region in Ig isotype-switched B cells. A mechanism for isotype stabilization.

Analysis of the Ig switch (S) region structure from many stable isotype-switched B cells reveals that the majority of the ultimate switch recombination sites are located at the 5' end, or even upstream of the 5' end of the Smu region. These findings strongly contrast with results from switch circle analysis which reveal that the primary switch recombination sites are preferentially distributed in the middle or at the 3' end of the Smu region. Secondary deletion within recombined chimeric S regions has been proposed as a mechanism to account for the different results. We directly tested whether secondary deletion of rearranged chimeric S regions indeed occurs in human cells. Circular DNAs representing secondary deletion events in gamma switched cells were isolated and characterized by using a specially designed PCR-based approach. Nucleotide sequence analysis revealed that all clones had the S gamma 1-Smu-S gamma 1 structure. Thus, these cloned fragments resulted from secondary deletion/recombination events within a chimeric Smu-S gamma 1 switch region, i.e., rearrangement between its 5' Smu and 3' S gamma 1. Analysis of the Smu region from stimulated B cells also revealed that some Smu regions undergo an internal deletion/rearrangement between the 5' and 3' ends. These results definitively demonstrate that secondary deletion/recombination of chimeric S regions of isotype-switched B cells occurs. Such secondary deletion/recombination events potentially can be responsible for isotype stabilization of switched B cells, as the active Ig gene may have an insufficient amount of retained chimeric switch sequence to serve as a substrate for further S-S recombination.