Regulation Of Class Switching Research Articles

Developmental and molecular regulation of immunoglobulin class switch recombination.

The ability of B lymphocytes to perform immunoglobulin (lg) class switch recombination depends largely on their differentiation stage. Pro-B cells, pre-B cells, and plasma cells do not switch at detectable levels. B cell receptor (BCR) cross linking, T cell help, and cytokines can drive the mature B cells into proliferation and differentiation and induce switch recombination. This recombination is targeted to distinct switch regions by cytokines through the induction of switch transcripts (Lorenz et al. 1995a, b). Cytokines also control the frequency of detectable, switched cells by regulating the amount of Ig secreted and the clone size of switched cells (for review, see Vercelli and Geha 1992; Coffman et al. 1993; Harriman et al. 1993; Lorenz and Radbruch 1996). Here, we focus on those molecules, which are involved in the regulation of class switching.

S region transcription per se promotes basal IgE class switch recombination but additional factors regulate the efficiency of the process.

Stimulation of B lymphocytes with a combination of lipopolysaccharide (LPS) and interleukin-4 (IL-4) induces germline transcription of and subsequent switching to the epsilon heavy chain constant region (C epsilon) gene. Mature germline C epsilon transcripts contain a non-coding exon (I epsilon exon) spliced to the C epsilon exons. To distinguish between the potential roles of germline transcription and those of germline transcripts in regulating the class switch process, we replaced the LPS- and IL-4-inducible I epsilon promoter and exon in ES cells with an LPS-inducible E mu enhancer/VH promoter expression cassette. Wildtype, heterozygous or homozygous mutant ES cells were injected into RAG-2 deficient blastocysts to generate somatic chimeras in which all B cells derived from ES cells. In contrast to normal B cells, heterozygous and homozygous mutant B cells had substantial transcription through the epsilon switch recombination region (S epsilon) following treatment with LPS alone and, under these conditions, both underwent low level switching (10- to 100-fold less than wildtype cells stimulated with LPS + IL-4) to IgE production. Heterozygous mutant cells underwent switching to IgE at essentially wildtype levels when stimulated with LPS and IL-4. However, homozygous mutant cells still showed extremely low levels of switching to IgE upon LPS and IL-4 stimulation. Analyses of hybridomas from heterozygous mutants indicated that the mutation is cis-acting and normal switching to other isotypes indicated that it is specific for IgE. Thus transcription per se generates low levels of class switch recombination in the absence of I region sequences. However, we demonstrate for the first time that, for optimal efficiency, the process requires the presence of the intact I region and/or I region promoter in cis, implicating factors beyond transcription through the S region in the regulation of class switching.

Replacement of germ-line epsilon promoter by gene targeting alters control of immunoglobulin heavy chain class switching.

Recent work has shown that the ability of cytokines to direct immunoglobulin heavy chain class-switch recombination to particular heavy chain constant (C) region (CH) genes correlates with the induction of specific germ-line CH transcripts. To test the role of germ-line transcripts in class switching, we have used homologous recombination to mutate the immunoglobulin heavy chain locus of the 18.81A20 murine pre-B-cell line. In the parent cell line, the combination of interleukin-4 (IL-4) and lipopolysaccharide (LPS) induces germ-line epsilon locus transcription prior to class switching to epsilon. The heavy chain locus of the mutated cell line contains the immunoglobulin heavy chain enhancer and variable region gene promoter in place of the LPS/IL-4-responsive germ-line epsilon promoter. The mutant cell line constitutively transcribes the epsilon locus in the absence of IL-4. Strikingly, the mutant cell line also switches to epsilon in the absence of IL-4. This result demonstrates that, at least in the 18.81A20 cell line, germ-line epsilon transcription plays a direct role in class switching to the epsilon locus. In addition, the ability to change the pattern of class switching by altering transcriptional activity indicates that transcription of germ-line CH is mechanistically important in regulation of class switching.