Growth of cell suspension cultures of tomato, Lycopersicon esculentum Mill. cv VFNT-Cherry, in the presence of cadmium is inhibited by buthionine sulfoximine, an inhibitor of glutathione synthesis. Cell growth and phytochelatin synthesis are restored to cells treated with buthionine sulfoximine by the addition of glutathione to the medium. Glutathione stimulates the accumulation of phytochelatins in cadmium treated cells, indicating that availability of glutathione can limit synthesis of these peptides.Exogenous glutathione causes a disproportionate increase in the level of smaller phytochelatins, notably [-y-Glu-CysJ2-Gly. In the presence of buthionine sulfoximine and glutathione, phytochelatins that are produced upon exposure to cadmium incorporate little [35SJcysteine, indicating that these peptides are probably not synthesized by sequential addition of cysteine and glutamate to glutathione.Plant cells that are exposed to cadmium rapidly accumulate peptides that have the general structure (y-Glu-Cys)n-Gly, where n = 2 to 10 (5, 11, 17). These peptides, variously termed PCs,4 poly(y-glutamyl-cysteinyl)glycines, y-glutamyl metal binding peptides, and cadystins, form intracellular complexes with cadmium. PCs have been identified in a large number of plant species (3) and also in some fungi, including Schizosaccharomyces pombe (12). The y-glutamyl linkages present in these peptides indicate that PCs are not direct translation products of mRNAs. However, the similarity of PCs to GSH (containing a single y-Glu-Cys moiety) suggests that GSH may be involved in the synthesis of PCs, and a number of observations support this hypothesis. Plant species that contain homo-GSH, with a carboxy terminal j3-alanine rather than glycine, synthesize homo-PCs that also contain a carboxy terminal ,3-alanine (6). When PC synthesis is induced by cadmium there is a rapid decline in cellular GSH levels (7, 16). Inhibition of 7y-Glu-Cys synthetase, the penultimate enzyme of the GSH synthesis pathway, by BSO prevents the accumulation of PCs (7,(15)(16)(17) MATERIALS AND METHODS Cell CulturesCell suspension cultures of tomato Lycopersicon esculentum Mill. cv VFNT-Cherry were maintained as described (16). Cultures were initiated with an inoculum of 20 mg cells (fresh weight) per mL of medium and grown with shaking at 24 to 260 C. Cells were subcultured weekly and experiments performed 3 or 4 d after inoculation. GSH, BSO, and CdC12 (Sigma Chemical Co.) were added to the medium as filter sterilized solutions. Assays for GSH and PhytochelatinsCells to be assayed for GSH and PCs were collected by vacuum filtration on Whatman No. 4 paper and frozen at -70°C. Extracts were prepared by adding an equal volume (1 mL per g fresh weight of cells) of 10% (w/v) 5-sulfosalicylic acid and keeping the mixture on ice for 10 min, during which the extracts were vortexed three times. The lysate was centrifuged at 13,000 g at 4°C for 4 min, and the acid soluble supernatant was either assayed immediately or stored at -70°C for future analysis. GSH and...

Listeria monocytogenes is a pathogenic bacterium that can grow at low temperatures and elevated osmolarity. The organism survives these stresses by the intracellular accumulation of osmolytes: low-molecular-weight organic compounds which exert a counterbalancing force. The primary osmolyte in L. monocytogenes is glycine betaine, which is accumulated from the environment via two transport systems: glycine betaine porter I, an Na ؉ -glycine betaine symporter; and glycine betaine porter II, an ATP-dependent transporter. The biochemical characteristics of glycine betaine porter I were investigated in a mutant strain (LTG59) lacking the ATPdependent transporter. At 4% NaCl, glycine betaine uptake in LTG59 was about fivefold lower than in strain DP-L1044, which has both transporters, indicating that the ATP-dependent transporter is the primary means by which glycine betaine enters the cell. In the absence of osmotic stress, cold-activated uptake by both transporters was most rapid between 7 and 12°C, but a larger fraction of the total uptake was via the ATP-dependent transporter than was observed under salt-stressed conditions. Twelve glycine betaine analogs were tested for their ability to inhibit glycine betaine uptake and growth of stressed cultures. Carnitine, dimethylglycine, and ␥-butyrobetaine appear to inhibit the ATP-dependent transporter, while trigonelline and triethylglycine primarily inhibit glycine betaine porter I. Triethylglycine was also able to retard the growth of osmotically stressed L. monocytogenes grown in the presence of glycine betaine.

The USDA germplasm repositories help to preserve the genetic variability of important crop species by collecting and maintaining representative cultivars and related germplasm. Simple sequence repeat markers with high allelic diversity were used to type 41 grapevines from 40 accessions. All vines were either seedless table grape cultivars or cultivars with names similar to table grape cultivars. The proportion of shared alleles was selected as the most appropriate statistical measure of genetic distance for this population. In conjunction with morphological traits, known synonyms were confirmed and a previously unknown synonym was discovered. An alleged synonym in the literature was disproved by the DNA data. The data were consistent with known parentage, where such data were available. Two mislabeled vines in the USDA collection were identified. UPGMA grouped the cultivars loosely into three groups: a group of nine mostly Middle Eastern cultivars, a group of 22 accessions mostly from Russia and Afghanistan that were morphologically similar to 'Thompson Seedless', and a third very loose group of 11 accessions consisting mostly of eastern European wine grape cultivars. The limitations and usefulness of this type of analysis are discussed.

Hybridization of introduced domesticates and closely related natives is well documented in annual crops. The widespread introduction of the domesticated grapevine, Vitis vinifera, into California where it overlaps with two native congenerics, with which it is interfertile, provides opportunity to investigate hybridization between woody perennials. Although geographically widespread, the introduction over the past two centuries has been limited to a few elite clonal cultivars, providing a unique opportunity to study the effects of hybridization on the native species. The amount of hybridization with V. vinifera and the genetic diversity of wild‐growing Vitis californica and Vitis girdiana were examined using nineteen microsatellite markers. STRUCTURE analysis was used to define hybrid and introgressed individuals and to analyze genetic structure of the native species. FAMOZ software was used to identify which V. vinifera cultivars served as parents of F 1 hybrids. The three species were clearly distinguished by STRUCTURE analysis. Thirty percent of 119 V. californica vines were hybrids. The domesticated parent was identified for 16 F 1 hybrid vines; the original California cultivar, ‘Mission’, was the parent of eight. Backcrosses were also found, showing introgression into subsequent generations. Similar results were obtained for a small sample of V. girdiana. Removing hybrids greatly reduced the genetic variation of the presumed pure species, among which there was essentially no genetic structure. Limited genetic variability indicates the California natives may be threatened by genetic erosion. The discovery of F 1 hybrids of ‘Mission’, a cultivar not grown in the areas for ~100 years, suggests long generation times for wild vines that, often, grow into expansive liana and propagate by layering, all factors that limit recruitment in populations already disjunct by habitat lose. Hermaphroditic flowers and fruit that is more attractive to birds may favor the production of backcross seed and establishment of introgressed individuals.

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