Effect of cytokines on switching to IgA and alpha germline transcripts in the B lymphoma I.29 mu. Transforming growth factor-beta activates transcription of the unrearranged C alpha gene.

Abstract

H chain isotype switch recombination is preceded by the appearance of RNA initiating 5' of the specific switch region that will undergo recombination. In an effort to understand the potential function of germline transcripts in switch recombination and whether the regulation of germline transcripts correlates with the regulation of switching, we are studying this process in the murine B lymphoma cell line I.29 mu, which switches, after treatment with bacterial LPS, primarily to IgA and less frequently to IgE. Levels of alpha germline transcripts initiating upstream of alpha switch (S alpha) sequences are elevated in clones of this line that switch well, compared with clones that switch less frequently. Transforming growth factor-beta (TGF-beta) has been shown to increase alpha germline transcripts and switching to IgA expression in LPS-stimulated murine splenic B cells. We now demonstrate that TGF-beta increases LPS-induced switching to IgA by 10-fold at optimal doses and increases the level of alpha germline transcripts 5- to 9-fold in I.29 mu cells. Nuclear run-on analysis shows that this increase is at the level of transcription. Thus, TGF-beta appears to direct switching to IgA by inducing transcription from the unrearranged S alpha-C alpha DNA segment. Germline alpha RNA is quite stable in I.29 mu cells, having a half-life of about 5 h, and we find no evidence for further stabilization in the presence of TGF-beta. Levels of epsilon germline transcripts are decreased by TGF-beta treatment. IL-4, which modestly increases switching in I.29 mu cells, slightly increases transcription of alpha germline RNA. IFN-gamma, which reduces switching to IgA in these cells, also reduces the level of alpha germline transcripts. IFN-gamma also reduces the level of epsilon germline transcripts induced by IL-4. Our results support the hypothesis that the regulation of transcription of particular switch sequences by cytokines regulates the specificity of recombination. We also present evidence that IL-4 may provide other signals, distinct from transcriptional targeting, that increase LPS-induced switching to IgA.