Treatment of human splenic B lymphocytes with the mitogen Branhamella catarrhalis (BC) and transforming growth factor-beta 1 (TGF-beta 1) induces expression of germline Ig C alpha transcripts and class switching to this isotype. To further characterize the molecular mechanism by which TGF-beta 1 and mitogenic signals regulate the expression of unrearranged C alpha 1 and C alpha 2 genes, we have characterized the promoter elements that are responsible for the transcriptional activation of their corresponding germline genes using transient expression assays. We report here that both in the I alpha 1 and the I alpha 2 regions, maximal phorbol myristate acetate (PMA) and TGF-beta 1 responsiveness of the promoters can be conferred by 327 bp spanning the transcription initiation sites and a previously identified phylogenetically conserved region. The expression of these 327 bp segments is not restricted to the B cell lineage since they are also active in the erythroleukemia cell line K562 as well as the B cell lines Raji and DG75. Mutational analyses have demonstrated the importance of sequences within the 327 bp segment that contain a putative cyclic AMP responsive element binding protein (CREB) binding site for TGF-beta 1 and PMA responsiveness and putative PU-1 and Sp1 binding sites for basal promoter activity. Upstream distal elements that could negatively modulate the expression of the I alpha 1 and I alpha 2 promoters, particularly in non-B cells, have been identified. Three such elements were mapped between positions -352 to -243, -627 to -516, and upstream of position -731 respectively. The influence of these elements presumably contributes to the B cell specific expression of the I alpha 1 and I alpha 2 promoters. The I alpha 1 and I alpha 2 promoters were found to be functionally indistinguishable from each other with respect to their basal level of expression, and their responsiveness to TGF-beta 1.