The tonoplast plays a crucial role in ion compartmentation, which is a central feature of the salt tolerance of halophytes, but we do not know the properties of the membrane that confer this ability. A method was, therefore, developed for the isolation of vacuoles from Suaeda maritima (L.) Dum. of sufficiently high purity to enable biochemical characterization of their lipid and protein composition. Tonoplast fractions produced by density gradient centrifugation, as well as vacuoles isolated by a variety of methods (including DEAE dextran lysis, digitonin lysis, and mechanical shear forces) were unacceptably contaminated. A highly pure vacuole preparation was obtained when protoplasts were lysed by a mild hypotonic shock in alkaline buffer, in the presence of the compatible cytosolute glycine-betaine, followed by shear force during ultracentrifugation; cytoplasmic contamination was prevented by the addition of the zwitterionic detergent 3-([3-cholamidopropyl]dimethylammonio)-l-propanesulphonate (CHAPS). Light microscopy of this preparation revealed no intact protoplasts and no contamination by chlorophyll could be detected. Electron microscopy showed the vacuoles to be single-membrane-bound structures, and was the only criterion upon which vacuoles could be separated reliably from vacuoplasts, in which the plasmalemma is collapsed on to the tonoplast. Analysis by SDS-PAGE showed that a total of 15 polypeptides were enriched in the tonoplast and 27 in the soluble fraction from vacuole preparations, with a pattern similar to that reported for glycophytic species. The pure tonoplast exhibited both vanadate-insensitive ATPase and pyrophosphatase activities, but the properties of these enzymes were broadly similar to those of glycophytes. Analysis of membrane fatty acids showed that the degree of saturation of the putative tonoplast preparation increased as the assessment of the purity of the preparation (made by microscopy) increased. The ATPase could be substantially purified by ion-exchange FPLC. The results are discussed in relation to the degree of purity needed in membrane preparations in order to be suitable for biochemical analysis.

While members of the genus Oryza are very sensitive to salinity, salt concentrations as high as 20% of that of seawater had no adverse effect on the growth of the tropical wild rice (Porteresia coarctata Tateoka) in experiments undertaken in a greenhouse in the UK. P. coarctata plants accumulated sodium and chloride ions in the leaves, but maintained a Na: K ratio as low as 0.7, even after 6 weeks of growth in 25% artificial seawater (ASW) where the Na: K ratio was 34. This ability to maintain a high K: Na ratio in the leaves is in part a consequence of the secretion of ions from the leaves. The ratio of Na: K in the secreted salt (more than 5 for plants growing in 25% ASW) is similar to that measured by X-ray microanalysis in the vacuoies of hairs found in folds of the adaxial surface of the leaf lamina, suggesting that the secretions emanate from these hairs. The salt secreted by the hairs is an important factor in the salt-balance of the leaves: the consequences of these findings for the transfer of salt-tolerance from this species into cultivated rice are discussed.

Vacuoles of high purity were isolated from the leaves of the halophyte Suaeda maritima (L.) Dum. The relative compositions of phospholipids, phytosterols, and fatty acids in the tonoplast membrane were determined and membrane fluidity was assessed by electron spin resonance. The characteristics of the tonoplast were consistent with minimizing passive permeability to NaCI. The phospholipid: protein ratio (1.1: 1.0) was higher than that recorded in other membrane preparations, including vacuoles from beetroot storage material, commensurate with the low density (1.05 g cm−3) of the S. maritima tonoplast. The tonoplast fatty acids were highly saturated and dominated by n-hexadecanoic acid and n-octadecanoic acid. Phytosterols identified by gas chromatography were cholesterol, campesterol, stigmasterol, and β-sitosterol. Cholesterol was a trace percentage in protoplasts, but comprised 30% of the tonoplast sterols. Semi-quantitative analysis by chromatography on silica gel revealed an enrichment in the tonoplast of glycolipid which was not accounted for as chloroplast contamination. The fluidity of the tonoplast, determined by electron spin resonance, was lower than the protoplasts, consistent with the high degree of saturation of the fatty acid chains. The relevance of the lipid composition of the tonoplast to its central role in ion compartmentation within the halophyte cell is discussed.

Phenotypic resistance of salinity is expressed as the ability to survive and grow in a salinised medium. Some subjective measure of overall performance has normally been used in plant breeding programmes aimed at increasing salinity resistance, not only to evaluate progeny, but to select parents. Salinity resistance has, at least implicitly, been treated as a single trait. Physiological studies of rice suggest that a range of characteristics (such as low shoot sodium concentration, compartmentation of salt in older rather than younger leaves, tolerance to salt within leaves and plant vigour) would increase the ability of the plant to cope with salinity. We describe the screening of a large number of rice genotypes for overall performance (using an objective measure based on survival) and for the aforementioned physiological traits. There was wide variation in all the characters studied, but only vigour was strongly correlated with survival. Shoot sodium concentration, which a priori is expected to be important, accounted for only a small proportion of the variability in the survival of salinity. Tissue tolerance (the cellular component of resistance reflecting the ability to compartmentalise salt within leaves) revealed a fivefold range between genotypes in the tolerance of their leaves to salt, but this was not correlated positively with survival. On the basis of such (lack of) correlation, these traits would be rejected in normal plant breeding practice, but we discuss the fallacies involved in attempting correlation between individual traits and the overall performance of a salt-sensitive species in saline conditions. We conclude that whilst overall performance (survival) can be used to evaluate the salt resistance of a genotype, it is not the basis on which parents should be selected to construct a complex character through breeding. It was the norm for varieties which had one good characteristic affecting salt resistance to be unexceptional or poor in the others. This constitutes experimental evidence that the potential for salt resistance present in the rice genome has not been realised in genotypes currently extant. The results are discussed in relation to the use of physiological traits in plant breeding, with particular reference to environmental stresses that do not affect a significant part of a species' ecological range.