Rapid decline thymidine kinase activity during differentiation of Naegleria.

Abstract

A logarithmically growing population of Naegleria gruberi differentiate into flagellates synchronously in about 85 to 90 minutes after the stimulation. Using this system, two separate enzymes that phosphorylate thymidine were assayed. Thymidine kinase activity decreases rapidly after the stimulation, while nucleoside phosphotransferase activity remains approximately constant. The possibility that this observed decrease in thymidine kinase activity is due to an expanded thymidine pool or to an increased phosphatase activity has been ruled out. Mixing experiments indicated that there is neither an activator of thymidine kinase in growing cells nor an inhibitor in differentiating cells. Thus, the observed decrease is likely to be due to an inactivation of the enzyme. Addition of actinomycin D or cycloheximide does not prevent the decrease in thymidine kinase activity suggesting that neither transcription nor translation is involved in the putative inactivation of thymidine kinase enzyme. There is very good correlation between the degree (or rate) of decrease in thymidine kinase activity and the percent of cells differentiating into flagellates. Thus, it appears certain that the residual thymidine kinase activity is primarily due to some portion of cells which did not respond to the stimulation and did not differentiate. Since there is an early termination of nuclear DNA synthesis in differentiating Naegeria, the decrease in thymidine activity (but no change in nucleoside phosphotransferase) suggests a correlation between thymidine kinase and DNA synthesis in Naegleria.