The role of zinc ions in the transformation of lymphocytes by phytohaemagglutinin

Abstract

1. Incorporation of [3H]thymidine into DNA was inhibited by EDTA and diethylenetriamine-NNN′N″N″-penta-acetate but not by nitrilotriacetate even when Ca2+ was present at more than twice the concentration of the chelators. 2. The inhibition caused by EDTA was most effectively reversed by Zn2+ but also to a lesser extent by Cd2+. Very little if any activation of the EDTA-inhibited system was obtained with Co2+, Cu2+, Fe3+, Mn2+ or Ni2+ added alone. 3. Fe3+ added to the Zn2+-activated lymphocytes in the presence of EDTA markedly increased thymidine incorporation. Addition of Cd2+ prevented the inhibition of incorporation which occurred at high Zn2+ concentrations. 4. If EDTA was added more than 15h after phytohaemagglutinin, the inhibition of incorporation was less than that obtained by its addition at zero time. If Zn2+ was added later than 12h after EDTA and phytohaemagglutinin, the inhibition of incorporation by EDTA was not fully reversed. A study of the time-course of the effects of delayed additions of EDTA and Zn2+ suggested that, on average, the cells required Zn2+ between 20 and 30h after phytohaemagglutinin addition to allow the full rate of thymidine incorporation at 37h. 5. The increase in the rate of protein synthesis caused by phytohaemagglutinin was not inhibited by EDTA until about 8h. The further increase after this was totally inhibited by EDTA but this inhibition was fully reversible with Zn2+. The rate of protein synthesis in EDTA-inhibited cultures at 40h was the same as that at 10h. 6. There was a close similarity between the effects of EDTA on lymphocyte stimulation and those reported by Kay et al. (1969) with low doses of actinomycin D.