Switch recombination in a transfected plasmid occurs preferentially in a Bcell line that undergoes switch recombination of its chromosomal Ig heavy chain genes

Ab class switching is induced upon B cell activation in vivo by immunizatio n or infection or in vitro by treatment,pith mitogens, e.g. LPS, and result s in the expression of different heavy chain constant region (C-H) genes wi thout a change in the Ab variable region. This DNA recombination event allo ws Abs to alter their biological activity while maintaining their antigenic specificity. Little is known about the molecular mechanism of switch recom bination, To attempt to develop an assay for enzymes, DNA binding proteins, and DNA sequences that mediate switch recombination, we have constructed a plasmid DNA substrate that will undergo switch recombination upon stable t ransfection into the surface IgM(+) B cell line (I.29 mu), a cell line capa ble of undergoing switch recombination of its endogenous genes. We demonstr ate that recombination occurs between the two switch regions of the plasmid , as assayed by PCRs across the integrated plasmid switch regions, followed by Southern blot hybridization, Nucleotide sequence analysis of the PCR pr oducts confirmed the occurrence of S mu-S alpha recombination in the plasmi d. Recombination of the plasmid in I.29 mu cells does not require treatment with inducers of switch recombination, suggesting that recombinase activit y is constitutive in I.29 mu. cells. Recombination does not require high le vels of transcription across the snitch regions of the plasmid, Fewer recom bination events are detected in four different B and T cell lines that do n ot undergo switch recombination of their endogenous genes.