PARTICIPATION OF CAMP IN THE INDUCTION OF THE SYNTHESIS OF GLYCEROL FOR THE OSMOREGULATORY RESPONSE IN THE SALT-TOLERANT YEAST ZYGOSACCHAROMYCES-ROUXII

The possible participation of cyclic AMP (cAMP) in osmoregulation during the initial stages of salt stress was examined in cells of a salt-tolerant yeast, Zygosaccharomyces rouxii. When the cells were exposed to a medium with 1 M NaCl, levels of cAMP reached a maximum within 10 min and then declined. The maximum level was about 4.6 times higher than the initial intracellular level, measured when the cells were inoculated into the medium. Adenylate cyclase (AC) activity, which catalyzes the synthesis of cAMP from ATP, increased immediately after exposure of cells to medium with 1 M NaCl, reached a maximum of 4.43 pmol cAMP produced/min.mg protein at 15 min and then decreased. The activity of phosphodiesterase (PDE), which catalyzes the conversion of cAMP to 5'-AMP, increased slowly after salt stress and reached a maximum at 25 min. The increase in PDE activity resulted in a decrease in the cAMP content of cells. These observations indicate that the levels of cAMP during salt stress are controlled by the activation and inactivation of the two enzymes. When GTP gamma S was added at 10 mu M to medium with 1 M NaCl, AC activity increased to about 2.5 times that in the case of medium with 1 M NaCl but without GTP gamma S. This result suggests that AC activity may be regulated by a GTP-binding regulatory protein (G-protein) during salt stress. When 2-deoxy-D-glucose (DG), an inhibitor of AC, was added at 8 mM to the medium with 1 M NaCl, synthesis of cAMP and AC activity were completely inhibited. Both the accumulation of glycerol and activation of the plasma membrane ATPase, induced by salt stress, were simultaneously inhibited in the presence of DG. This result indicates that the increase in the cAMP content of cells was related to the intracellular accumulation of glycerol and to activation of a plasma membrane ATPase during salt stress. These results together suggest that cAMP synthesized under regulatory control exerted by a G-protein may function as a second messenger in osmoregulation in response to salt stress.