On the role of phosphate and phosphoprotein in thymocyte interphase death

Abstract

According to Whitfield's hypothesis, interphase death in thymocytes results from accumulation of phosphate and from increased phosphoprotein synthesis in the nucleus which is caused by an inhibition of nuclear oxidative phosphorylation. The experiments described in this paper provided no evidence either for phosphate accumulation or for increased protein phosphorylation under the conditions predicted by the hypothesis. One and two hours after irradiation with 800 rads and incubation in a medium containing 30 mM phosphate, the intracellular phosphate content and the amount of protein-bound phosphate had not changed. Also the exchange of phosphate with the medium had not changed while the labelling of protein-bound phosphate seemed to be slightly inhibited. Addition of 5 × 10 -4 M dinitrophenol after irradiation decreased the appearance of pyknotic nuclei and strongly increased the intracellular phosphate content but had no effect on the amount or labelling of protein-bound phosphate. Oligomycin did not influence phosphoprotein labelling, it inhibited phosphate exchange between cells and medium but did not increase pyknosis; on the contrary it inhibited radiation-induced pyknosis to a greater extent than did dinitrophenol. Cortisol increased pyknosis without any effect on phosphate and phosphoprotein, and this increase was inhibited by dinitrophenol. Valinomycin had no inhibitory effect on nuclear phosphorylation and did not increase intracellular phosphate and phosphoprotein; it did increase the rate of pyknosis. We conclude that the mechanism initiating interphase death in irradiated thymocytes remains unknown.