Effects of cycloheximide and actinomycin D on the amino acid transport system of tetrahymena

Abstract

Tetrahymena pyriformis were grown in axenic culture to late logarithmic and stationary phases, resuspended in an inorganic medium, and the rates of transport of alpha-aminoisobutyric acid (AIB) and of the decarboxylation of L-[1-14C]leucine and L-[1-14C]tyrosine were measured. There was a rapid loss of each of these measures of amino acid transport in both late log phase and stationary phase cells, Addition of actinomycin D to the washed cells caused a small increase in the rate of loss of capacity to decarboxylate tyrosine and leucine. Addition of cycloheximide to the washed cells caused a reduction in the rates of loss of capacity to transport AIB and to decarboxylate leucine and tyrosine except that in late log phase cells cycloheximide markedly increased the rate of loss of capacity to decarboxylate leucine. When cells that had been pretreated with chlorpromazine to reduce their amino acid transport capacity were washed and resuspended in proteose peptone the capacity to decarboxylate tyrosine and leucine increased to control values within 1.5 hours. Addition of actinomycin D reduced the rate of recovery of transport systems in this cell. The finding that AIB and N-methylaminoisobutyrate are both taken up by Tetrahymena, the latter at one-eighth the rate of the former, but that neither one alters the rate of uptake of the other provides preliminary support for this possibility. The present results further suggest that the transport system(s) has a short lifetime and that the balance between rate of synthesis and rate of loss of the transport system is controlled in part by the presence of exogenous amino acids.

Regulation of taurine accumulation in the ciliate protozoanTetrahymena pyriformis

“…The observations presented in Table 1 support the notion by Blum (1982) that in the absence of any transportable amino acids, Tetrahymena not only stops or greatly reduces the rate of synthesis of its amino acid transporting systems but it also seems to rapidly inactivate or degrade these systems. The data in Fig.…”

“…4B). Assuming that protease activity is a prerequisite for protein degradation leading to the loss of amino acid transport capacity, this activity is insensitive to leupeptin and pepstatin according to Blum (1982). PPY itself is a protease inhibitor (Jonasssen and Grinde, 1986) and the gradual loss of taurine transport capacity seen after transfer from PPY to inorganic medium is, therefore, likely to result from an increased action of proteases on the transport systems.…”

“…Tetrahymena accumulates various amino acids against a concentration gradient and it appears that most if not all of the transport occurs across the plasma membrane via distinct amino acid transporting systems and not by endocytosis (Hoffmann and Rasmussen, 1972;Blum, 1982;Stephens, 1983, 1986; see Dunham and Kropp, 1973;Blum, 1982). The influx of certain amino acids in Tetrahymena is affected by the presence of other amino acids in the extracellular compartment (see Dunham and Kropp, 1973) and besides the taurine transporter, Tetrahymena also has two other amino acid transporting systems which can be distinguished by their ability to transport AIB and meAIB (Blum, 1982).…”

“…The influx of certain amino acids in Tetrahymena is affected by the presence of other amino acids in the extracellular compartment (see Dunham and Kropp, 1973) and besides the taurine transporter, Tetrahymena also has two other amino acid transporting systems which can be distinguished by their ability to transport AIB and meAIB (Blum, 1982). However, even though influx of free amino acids into Tetrahymena might only play a minor role to the contribution of nutrients (Davis and Stephens, 1983) the cells lose their capacity to take up mnino acids following transfer to a non-nutrient medium (Blum, 1982). Similarly, starvation of Tetrahymena causes a reduction in the initial taurine influx, i.e., a reduction in transport via the Na+-dependent taurine transporter as well as via the taurine leak pathway (Table 1).…”

“…It has previously been shown that Tetrahyrnena pyriforrnis is able to accumulate the non-metabolizable amino acid AIB against a concentration gradient and that this capacity is gradually reduced after transfer of the cells to a nonnutrient medium (Blum, 1982). The taurine influx in Tetrahyrnena was accordingly estimated at various time points after transfer of the cells from the PPY medium to taurine free IMP.…”