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Functional Analysis of I Promoter Regions of Multiple IgA Heavy Chain Genes¹

Department of Microbiology and Immunology, Stritch School of Medicine, Loyola University Chicago, Maywood, IL 60153

The 13 nonallelic IgA H chain genes of rabbit are differentially expressed in vivo. They can be grouped into those expressed at high levels (C4, C5, C6, C9, C10, C12, and C13), those expressed at low levels (C1, C2, C7, and C11), and those that are not expressed (C3 and C8). We tested whether the differential in vivo expression is due to differential responses of the I promoters to TGF- stimulation. We stimulated the rabbit B cell line 55D1 with TGF- and, using single-cell RT-PCR, found that expression of germline (GL) transcripts of 3 and 8 could not be induced. By luciferase reporter gene assay and EMSA we found that the promoters of the unexpressed isotypes C3 and C8 are defective, thereby explaining the absence of IgA3 and IgA8 in vivo. When comparing the promoter activities of the other isotypes we found that the activities did not reflect the degree of in vivo expression. Instead, the promoters of the isotypes expressed at high or low levels promoted expression of the luciferase gene to a similar degree, except for the I4 promoter, which had much higher activity. Also the degree to which TGF- induced GL expression of the various isotypes in 55D1 B cells did not reflect in vivo expression. However, most of the TGF--stimulated cells expressed GL mRNA of multiple isotypes; no isotype was expressed preferentially. These results suggest that the final switch to a single isotype is regulated in a step subsequent to GL transcription, rather than by induction of GL transcripts by the I promoter.

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