Enhanced Net K Uptake Capacity of NaCl-Adapted Cells.

Maintenance of intracellular K(+) concentrations that are not growth-limiting, in an environment of high Na(+), is characteristic of NaCl-adapted cells of the glycophyte, tobacco (Nicotiana tabacum/gossii). These cells exhibited a substantially greater uptake of (86)Rb(+) (i.e. an indicator of K(+)) relative to unadapted cells. Potassium uptake into NaCl-adapted cells was 1.5-fold greater than unadapted cells at 0 NaCl and 3.5-fold greater when cells were exposed to 160 millimolar NaCl. The difference in net K(+) uptake between unadapted and NaCl-adapted cells was due primarily to higher rates of entry rather than to reduced K(+) leakage. Presumably, enhanced K(+) uptake into adapted cells is a result of electrophoretic flux, and a component of uptake may be linked to vanadate-sensitive H(+) extrusion.

Abscisic Acid Stimulated Osmotic Adjustment and Its Involvement in Adaptation of Tobacco Cells to NaCl.

Osmotic adjustment of cultured tobacco (Nicotiana tabacum L. var Wisconsin 38) cells was stimulated by 10 micromolar (+/-) abscisic acid (ABA) during adaptation to water deficit imposed by various solutes including NaCl, KCl, K(2)SO(4), Na(2)SO(4), sucrose, mannitol, or glucose. The maximum difference in cell osmotic potential (Psipi) caused by ABA treatment during adaptation to 171 millimolar NaCl was about 6 to 7 bar. The cell Psipi differences elicited by ABA were not due to growth inhibition since ABA stimulated growth of cells in the presence of 171 millimolar NaCl. ABA caused a cell Psipi difference of about 1 to 2 bar in medium without added NaCl. Intracellular concentrations of Na(+), K(+), Cl(-), free amino acids, or organic acids could not account for the Psipi differences induced by ABA in NaCl treated cells. However, since growth of NaCl treated cells is more rapid in the presence of ABA than in its absence, greater accumulation of Na(+), K(+), and Cl(-) was necessary for ion pool maintenance. Higher intracellular sucrose and reducing sugar concentrations could account for the majority of the greater osmotic adjustment of ABA treated cells. More rapid accumulation of proline associated with ABA treatment was highly correlated with the effects of ABA on cell Psipi. These and other data indicate that the role of ABA in accelerating salt adaptation is not mediated by simply stimulating osmotic adjustment.

Proton Fluxes as a Response to External Salinity in Wild Type and NaCl-Adapted Nicotiana Cell Lines.

Addition of 100 millimolar KCl, NaCl, or Na(2)SO(4) strongly promoted acidification of the medium by cells of Nicotiana tabacum/gossii in suspension culture. Acidification was greater in the case of NaCl-adapted than in that of wild type cells, and strikingly so in KCl medium when fusicoccin (FC) was present. Back-titration indicated that net proton secretion in KCl medium was increased 4-fold by FC treatment in the case of adapted cells; but was not even doubled in wild type cells. Membrane potential was higher in NaCl-adapted cells. FC treatment hyperpolarized wild, but not NaCl-adapted cells, suggesting a higher degree of coupling between H(+) efflux and K(+) influx in adapted cells; FC enhanced net K(+) uptake in adapted but not in wild cells. Acidification by cells suspended in 10 millimolar KCl was highly sensitive to vanadate, but that after addition of 100 millimolar KCl or NaCl was much less sensitive. Addition of 100 millimolar NaCl to wild type cells already provided with 10 millimolar KCl briefly accelerated, then slowed down the rate of acidification. If the addition was made after acidification had already ceased, alkalization was observed, particularly in the presence of FC. The results are consistent with the operation of a Na(+)-H(+) antiporter.

Comparison between a Stable NaCl-Selected Nicotiana Cell Line and the Wild Type : K, Na, and Proline Pools as a Function of Salinity.

An NaCl-resistant line has been developed from suspension-cultured tobacco cells (Nicotiana tabacum/gossii) by stepwise increases in the NaCl concentration in the medium. Resistance showed stability through at least 24 generations in the absence of added NaCl.Above an external NaCl concentration of 35 millimolar, proline concentration in the selected cells rose steeply with external NaCl, particularly so above 100 millimolar NaCl. Proline accumulation in the wild type was far slighter. Selected cells which had been grown for 24 generations in the absence of added NaCl accumulated proline strongly on re-exposure to NaCl medium, indicating stability of this character. Proline accumulation was fully reversible with a half-time of about 6 hours. When selected cells were transferred sequentially to lower and lower NaCl concentrations, their proline content fell to the level corresponding to the new NaCl concentration. The NaCl-selected cells responded to water stress (i.e. added mannitol) by accumulating markedly more proline than did the wild type.The addition of Ca(2+) to the growing and rinsing media minimized Na(+) and K(+) binding in the Donnan free space of cell walls and thus allowed assessment of intracellular Na(+) and K(+). In both cell types, internal Na(+) content rose steadily as a function of external NaCl concentration. In the course of 7 days in NaCl media, the wild type cells lost a considerable part of their K(+) content, the extent of the loss increasing with rise in external NaCl concentration. The selected cells, by contrast, lost no K(+) at external NaCl concentrations below 50 millimolar external NaCl, and at higher concentrations lost less than the wild type.