Differential Regulation of Mouse Germline Ig γ1 and ε Promoters by IL-4 and CD40

Abstract

Before Ig class switching, RNA transcription through the specific S regions undergoing recombination is induced by cytokines and other activators that induce and direct switching. The resulting germline (GL) transcripts are essential for switch recombination. To understand the differential regulation of mouse IgG1 and IgE, we compared the promoters for GL γ1 and ε transcripts. We addressed the question of why the promoter that regulates GL ε transcription is more responsive to IL-4 than the γ1 promoter and also why GL ε transcription is more dependent on IL-4 than is γ1 transcription. We found that the IL-4-responsive region of the GL ε promoter is more inducible than that of the γ1 promoter, although each promoter contains a binding site for the IL-4-inducible transcription factor Stat6, located immediately adjacent to a binding site for a basic region leucine zipper (bZip) family protein. However, the arrangement and sequences of the sites differ between the ε and γ1 promoters. The GL ε promoter binds Stat6 with a 10-fold higher affinity than does the γ1 promoter. Furthermore, the bZip elements of the two promoters bind different transcription factors, as the GL ε promoter binds and is activated by AP-1, whereas the γ1 promoter binds and is activated by activating transcription factor 2. C/EBPβ and C/EBPγ also bind the γ1 bZip element, although they inhibit rather than activate transcription. However, inhibition of promoter activity by C/EBPβ does not require the bZip element and may instead occur via inhibiting the activity of NF-κB.

Control of the STAT6–BCL6 Antagonism by SWAP-70 Determines IgE Production

“…Association of STAT6 with Ig1 was generally undetectable, indicating a weak and/or transient interaction of STAT6 with the g1 promoter. This is consistent with earlier data demonstrating different binding affinities of STAT6 for the ε (high affinity) and g1 (low affinity) promoter (27). Control PCRs diagnosing promoter regions of IgG2a (Ig2a) and of a proteasome gene, (Pr) (21,22), which lack STAT6 binding sites, did not generate signals (Fig.…”

“…Indeed, IgE production is the most tightly regulated of all Ig classes, and despite overlapping induction pathways, different regulatory elements govern the switch to IgG1 and IgE (26,27). Generation of IgE and its GLT specifically require signaling from the IL-4R (2,7,8,27), whereas g1 GLT depends less on IL-4R signaling (27). Therefore, we initially hypothesized a function for SWAP-70 in IL-4 signaling.…”

“…Germline g1 transcripts, which appeared earlier than the ε GLT, were downregulated 2-to 4-fold, although production of IgG1 was only very mildly affected by SWAP-70 deficiency, possibly indicating a less stringent requirement for efficient GLT at the g1 locus. Indeed, IgE production is the most tightly regulated of all Ig classes, and despite overlapping induction pathways, different regulatory elements govern the switch to IgG1 and IgE (26,27). Generation of IgE and its GLT specifically require signaling from the IL-4R (2,7,8,27), whereas g1 GLT depends less on IL-4R signaling (27).…”

“…These data suggest weaker association of STAT6 to chromatin in the absence of SWAP-70 in stimulated and nonstimulated cells. Because STAT6 occupancy of the Iε promoter requires high-affinity binding (27), we hypothesized that in the absence of SWAP-70 STAT6 may be much reduced or absent from this promoter.…”

“…The presumably weaker association of STAT6 with chromatin in Swap-70 2/2 cells raises the question as to whether this correlates with poor binding to Iε and hence reduced Iε GLT and IgE production. STAT6 needs to bind with different affinity to different promoter regions, and the Iε requires high-affinity binding (27).…”