Effects of osmotic shock and triton treatment on sorbose transport in Neurospora

Abstract

1. Conidia of Neurospora crassa were either subjected to osmotic shock or treated with the detergent triton X-100. A decrease in the amount of 14C-sorbose taken up, as measured by means of the millipore filter technique, served as criterium for the efficiency of both procedures. Treated cells were either a) pregerminated with fructose, b) ungerminated but preincubated with sorbose or c) ungerminated and not preincubated, to check for a possible loss of fructose-inducible, sorbose-inducible or constitutive transport components respectively. 2. In contrast to reports for bacteria, osmotic shock by transferring conidia from 30 - 60% sucrose solutions to 5·10-4м MgCl2-solution after 5 minutes did not decrease transport activity. Rather there was an increase in the amount of sorbose taken up after such treatment (Table 1). However, if sucrose was replaced by concentrated minearl salts solution (Vogels minimal 4.75 м stock solution) a drastic decrease of sorbose uptake was observed in conidia pregerminated with fructose (Fig. 1). Ungerminated conidia preincubated with sorbose did not respond to a shock with mineral salts solution; they did however respond, if 2% triton was added to the salts solution (Table 2). Triton alone was uneffective under comparable conditions (Table 2), but it became effective if the triton concentration was increased and the duration of treatment was extended to 30 minutes (Fig. 2). Ungerminated, not preincubated conidia showed the same reduction of sorbose uptake if treated with triton in this way (Fig. 2). 3. Whenever a decrease of sorbose uptake was observed, it was correlated with a decrease in cell survival. However, the amount of killing does not suffice to explain in full the decrease in sorbose uptake per sample. Thus even surviving cells demonstrate a decrease of sorbose uptake, the reasons for which are considered as follows: a) Cells preloaded with 14C-sorbose show the same rate of sorbose efflux, whether shocked with mineral salts solution or not (Fig. 3). The shock therefore does not cause an unspecific leakiness of the cell wall or membrane. b) Oxygen consumption in cell suspensions, as measured by an oxygen macro electrode, is strongly retarded for fructose pregerminated cells which have been shocked with mineral salts solution (Fig. 4). This retardation together with killing is fully sufficient to explain the decrease in sorbose uptake observed. Ungerminated cells preincubated with sorbose do not show this effect, not even after prolonged treatment with triton, when their ability to take up sorbose is reduced. Hence, in such material a loss of specific transport components is indicated.