Studies of mechanisms of antibody formation: III. Effects of 5-bromodeoxyuridine upon normal and neoplastic B-lymphocyte function

Although 5-bromodeoxyuridine (BrdU) does not interfere with the rate of proliferation, or with total protein, DNA, or RNA synthesis in normal mouse spleen B cells exposed to specific antigenic excitation or to polyclonal activators, this agent has the capacity to block the appearance of IgM antibody-forming cells (AFC) in response to antigenic challenge in vitro with sheep red blood cells (sRBC), as well as the generation of anti-sRBC or 2,4,6-trinitrobenzene sulfonate-AFC in the course of polyclonal expansion. The effect of BrdU can be blocked competitively by a 10-fold excess of thymidine (TdR). CsCl density equilibrium analysis indicates that B cells with 55% BrdU substitution of TdR in a single DNA strand could participate in at least one additional division cycle, resulting in a double DNA strand substitution. BrdU substitution in a single DNA strand was sufficient, however, to block antibody secretion in B cells (unifilar dominance). When cells with single-strand BrdU substitution were allowed to proliferate in the presence of a 10-fold excess of TdR, the cells regained the capacity to synthesize and secrete specific antibody, demonstrating the reversibility of the inhibitory effect of BrdU upon this specialized B-cell function. Cells stimulated with LPS in the presence of BrdU exhibited higher rates of synthesis of total RNA as well as of the RNA species of the translation machinery (28, 18, and 5S and tRNA), measured by the rate of incorporation of radioactive precursors at the time of peak proliferation. Exposure to BrdU blocked the secretion of μ and light chains, although such cells had high levels of membrane IgM (mIgM) and were able to regenerate mIgM after enzymatic stripping. The regeneration of mIgM could be suppressed with α-amanitine and by actinomycin D, suggesting that the incorporation of BrdU into DNA does not affect the transcripition of nucleotide sequences for mIgM. MOPC 315 and 70 plasmocytoma cells cultured in the presence of BrdU for 60 hr regularly incorporated BrdU into one and two DNA strands. There was a 55–60% substitution of BrdU in single DNA strands. This situation produces complete inhibition of the production of IgM, μ, or light chains in normal cells. In contrast, plasmocytoma cells treated with BrdU secreted as many α, γ, and light chains as the corresponding control cells grown in the presence of TdR. This result points to possibly profound differences in the regulation of transcripition of Ig nucleotide sequences in lymphocytes which have undergone malignant transformation.