Correlation of decreased calcium contents with proline accumulation in the marine green macroalga Ulva fasciata exposed to elevated NaCl contents in seawater

The involvement of Na+, K+, Cl− or Ca2+ in the regulation of salinity stress-induced proline accumulation via the inhibition of the activity of proline dehydrogenase (PDH; EC 1.4.3.1), a catabolic enzyme of proline, was investigated in the marine green macroalga Ulva fasciata Delile. After 6 h of exposure to elevated artificial seawater (ASW) salinity, adjusted either by increasing the NaCl content in 30‰ ASW (a change in ion ratio) or by concentrating ASW (a constant ion ratio), the contents of Na+, K+ and Cl− linearly accumulated with increasing salinity from 30–90‰ (parts per thousand); the accumulation pattern of each ion was similar between the two treatments. An increase in NaCl content in ASW induced proline accumulation, but decreased both the PDH activity and the total and water-soluble Ca2+ contents, while concentrated ASW had no effect. As compared to a constant value at 30‰, both the contents of total and water-soluble Ca2+ and the activity of PDH decreased 1 h after exposure to 60‰ (adjusted by increasing NaCl content in 30‰ ASW) and concomitantly the content of seawater Ca2+ increased, while proline accumulated after 3 h. The addition of 15 mM ethylene glycol-bis(2-aminoethyl ether) N,N,N'-tetraacetic acid (EGTA) in 60‰ ASW (adjusted by increasing the NaCl content in 30‰ ASW) enhanced both the proline accumulation and the decrease in the content of total and watersoluble cellular Ca2+ and the activity of PDH; the effects of EGTA were reversed by 10 mM CaCl2 or 10 mM CaSO4. These results indicate that a loss of cellular Ca2+ is associated with the NaCl induction of proline accumulation via an inhibition of PDH activity in U. fasciata.