Physarum Thymidine Kinase A Step or a Peak Enzyme Depending upon Temperature of Growth

Abstract

The variations of thymidine kinase or ATP : thymidine 5'-phosphotransferase (EC 2.7.1.21) during the cell cycle of Physarum polycephalum plasmodia have been studied at two extreme physiological temperatures: 22'C and 32°C. At 22°C the enzyme activity increases near mitosis and stays constant during late S and G2 phases, exhibiting the typical pattern of a 'step enzyme'. But at 32 "C thymidine kinase activity goes through a maximum 1 h 30 min after mitosis and decreases during the subsequent phases as expected for a 'peak enzyme'. The rate of enzyme degradation andlor inactivation, measured in the presence of metabolic poisons (cycloheximide or dinitrophenol), appears to follow a simple exponential function with a half-life of approximately 3 h and 1 h at 22 "C and 32 "C respectively. The effect of growth temperature on the decrease of thymidine kinase activity can account entirely for the differences in the pattern of enzyme activity at the two extreme temperatures. Tentative calculations indicate that the rate of enzyme synthesis is nearly constant during the cell cycle except near mitosis, where it is temporarily increased. The results suggest the existence of a regulatory mechanism able to modulate the rate of synthesis of thymidine kinase during the cell cycle.

The Effect of Nocodazole on Cell Cycle Events in the Plasmodial Slime Mould Physarum Polycephalum

“…We did not find any evidence for a ring-like configuration of condensed chromatin or microtubule inclusions as reported by others after griseofulvin or nocodazole treatment (Gull & Trinci, 1974;Wright et al, 1976;Herbert et al, 1980), but this was probably due to the fact that in these latter reports nuclei were examined after much longer exposure to the drtqy. The observations reported here substantiate previous work which had indicated that only completion of the initial stages of mitosis are necessary to trigger DNA replication (Brewer & Rusch, 1968;Wright & Tollon, 1978;Wolf et al, 1979;Wright et al, 1984). Mitosis and preparation for DNA replication may therefore be triggered by the same mechanism.…”

“…From temperature-shift experiments, Wright & Tollon (1978, 1979b have suggested the existence of a heat-sensitive pathway which is necessary for triggering both the increase in TdR kinase activity and the onset of mitosis. Nocodazole may be interfering with the regulation of the same pathway of events.…”

“…The experiments of Guttes & Guttes (1968) indicated that D N A replication was triggered by passage of the nuclei through mitosis. However, only the occurrence of the early events in the mitotic process seemed necessary for the onset of D N A replication because if the terminal stages (anaphase and telophase) were prevented, by heat shock (Brewer & Rusch, 1968;Wright & Tollon, 1978;Wolf et a/., 1979) or by the action of the anti-tumour drug cysplatin (Wright eta/., 1984), D N A replication still occurred.…”

Induction of Heat-Shock Proteins at Permissive Growth Temperatures in the Plasmodium of the Myxomycete Physarum polycephalum

“…It is tempting to hypothesize that the increased synthesis of these proteins is correlated with the mitotic delays which are observed after some temperature shifts up between two permissive temperatures [5,7]. Likewise, the mitotic delays induced by heat shocks to non-permissive temperatures either in Physarum [8] or in other eucaryotic cells [28 -331 and the synchronizing effects which can be obtained by some heat-shock regimens [34-371 could be due to the same mechanism.…”

“…A transient inactivation of the triggering substance can account for the mitotic delays induced by temperature shifts to the highest permissive temperature [5,71. This effect could be explained in several ways : the preferential inactivation at 32 "C of the triggering substance which has been synthesized at 22 "C [5, 71; an arrest in protein synthesis leading to an inbalance between the synthesis and the inactivation of the triggering substance after a shift to 32 "C; an inactivation of the triggering substance caused by the transient synthesis of an inhibitor induced by temperature shifts [7]. In order to exclude or to favor one of these possibilities we investigated the synthesis of new proteins after temperature shifts.…”