The Effects of High Salinity, Water-Deficit, and Abscisic Acid on Phosphoeno/pyruvate Carboxylase Activity and Proline Accumulation in Mesembryanthemum crystallinum Cell Cultures

Abstract

It has been established from previous studies in the halophyte Mesembryanthemum crystallinum that saline conditions induce the synthesis of phospho enol pyruvate carboxylase (PEPC), the performance of Crassulacean acid metabolism (CAM), and the production of compatible solutes such as proline. The objective of the present study was to determine whether factors causing the induction of these processes in leaves were also effective in tissue cultures of M. crystallinum grown under different conditions. In heterotrophically (sucrose)-grown calli of M. crystallinum , low levels of NaCl (10 to 25 mM) were required for optimum growth, while increasing concentrations of salt up to 200 mM caused a progressive decline in growth. Addition of salt to the culture medium caused a progressive increase in proline accumulation (13 fold increase in calli grown in 200 mM NaCl compared with calli grown without salt) but had little affect on PEPC activity in calli grown in sucrose media in the dark or light. Addition of 10 -7 M abscisic acid in the presence of 200 mM NaCl doubled the proline content in light-grown calli, while abscisic acid alone had no affect. However, growth with 10 -7 to 10 -5 M abscisic acid had no affect on levels of PEPC in dark or light-grown calli, in contrast to the previously reported induction of this enzyme and CAM by provision of abscisic acid to whole plants. Water stress (imposed by addition of polyethylene glycol) and low levels of NaCl (10 mM) in combination caused a large increase in proline content in heterotrophically cultured cells, whereas neither were effective alone. In photomixotrophic cell cultures, obtained by replacement of sucrose by soluble starch as the external carbon supply, there was a much higher chlorophyll content, and a limited salt induced increase in PEPC activity (4 fold enhancement during a 12-d, 200-mM, salt-shock period).