EFFECTS OF OSMOTIC-STRESS AND GROWTH STAGE ON CELLULAR PH AND POLYPHOSPHATE METABOLISM IN NEUROSPORA-CRASSA AS STUDIED BY P-31 NUCLEAR-MAGNETIC-RESONANCE SPECTROSCOPY

High-resolution P-31-NMR was employed to investigate the effects of gr owth stage and environmental osmolarity on changes of polyphosphate me tabolism and intracellular pH in intact Neurospora crassa cells. Our s tudy showed that changes of these parameters were growth-dependent. Th e ratio of polyphosphate to orthophosphate in vacuoles increased from 2.4 to 13.5 in N. crassa as cells grew from early log phase to station ary phase. Cytoplasmic pH and vacuolar pH changed, respectively, from 6.91 and 6.49 in early log phase cells to 7.25 and 6.84 in stationary phase cells. Hypoosmotic shock of N. crassa produced growth-dependent changes including: (i) a rapid hydrolysis of polyphosphate with a conc omitant increase in the concentration of the cytoplasmic phosphate, (i i) an increase in cytoplasmic pH, and (iii) an increase in vacuolar pH . Early log phase cells produced the most dramatic response whereas th e stationary phase cells appeared to be recalcitrant to the osmotic st ress. Thus, 95% and 60% of polyphosphate in the early log phase and mi d-log phase cells, respectively, disppeared in response to hypoosmotic shock, but little or no hydrolysis of polyphosphate occurred in stati onary cells. The cytoplasmic pH and the vacuolar pH increased in respo nse to hypoosmotic shock by 0.4 and 0.53 unit, respectively, in early log phase cells; and by 0.22 and 0.27 unit, respectively, in the mid-l og phase cells. In contrast, hypoosmotic shock of the stationary phase cells did not cause any change in intracellular pH. The osmotic stres s-induced polyphosphate hydrolysis and pH changes in early log and mid -log phase cells were reversible, suggesting that these changes were r elated environment osmolarity. Addition of polyamines or basic amino a cids which are known to be sequestered in vacuoles did not affect poly phosphate metabolism.