Although a detailed picture is emerging about the nature of the second messengers involved in B cell activation and proliferation, little is yet known about the intracellular events taking place further downstream. The c-myb proto-oncogene, the structurally related B-myb gene, and c-myc probably code for transcription factors, have been demonstrated to be necessary for the proliferation of hemopoietic cells, and their expression is indeed induced after mitogenic stimulation of T and B lymphocytes. They are therefore likely to be key elements in the regulation of gene expression during proliferation. We have set out to study the regulation of the expression of these two myb genes and of that of c-myc in relation to entry into the different phases of the cell cycle during mitogenic stimulation of resting human B lymphocytes. Resting tonsillar B cells stimulated with the anti-CD20 antibody 1F5 alone are induced to enter the G1 but not the S phase of the cell cycle, whereas co-stimulation with the anti-CD40 antibody G28.5 further drives them to enter the S phase and proliferate. The G28.5 antibody alone has been reported to partially activate and increase the alertness of resting B cells without inducing them to enter G1. In this report we show that increasing the strength of the activating signal leads to progressive induction of the proliferation-related genes studied. Thus the G28.5 antibody alone induces c-myc mRNA only in resting B cells, 1F5 induces both c-myc and B-myb, and the full mitogenic signal given by both antibodies together is accompanied by increased expression of all three--c-myc, B-myb, and c-myb genes. In addition, using a semi-quantitative polymerase chain reaction method, we show that different inhibitors of B cell proliferation, namely, cyclosporin A, an anti-CD19 antibody (HD37), and transforming growth factor beta 1 (TGF-beta 1), inhibit differentially the induction of these same genes after mitogenic stimulation of B cells. Whereas cyclosporin A inhibits induction of all three genes, TGF-beta 1 specifically blocks B-myb induction and CD19 has little effect on either of the genes tested. We conclude that c-myb, B-myb, and c-myc are regulated independently from one another, that induction of c-myc and B-myb together is not sufficient to trigger B cell proliferation, and we suggest that expression of all three is a prerequisite for proliferation to occur.