A characteristic feature of grasses and commercially important cereals is the presence of (1,3;1,4)-beta-d-glucans in their cell walls. We have used comparative genomics to link a major quantitative trait locus for (1,3;1,4)-beta-d-glucan content in barley grain to a cluster of cellulose synthase-like CslF genes in rice. After insertion of rice CslF genes into Arabidopsis, we detected (1,3;1,4)-beta-d-glucan in walls of transgenic plants using specific monoclonal antibodies and enzymatic analysis. Because wild-type Arabidopsis does not contain CslF genes or have (1,3;1,4)-beta-d-glucans in its walls, these experiments provide direct, gain-of-function evidence for the participation of rice CslF genes in (1,3;1,4)-beta-d-glucan biosynthesis.

The walls of grasses and related members of the Poales are characterized by the presence of the polysaccharide (1,3, 1,4)-␤-Dglucan (␤-glucan). To date, only members of the grass-specific cellulose synthase-like F (CSLF) gene family have been implicated in its synthesis. Assuming that other grass-specific CSL genes also might encode synthases for this polysaccharide, we cloned HvC-SLH1, a CSLH gene from barley (Hordeum vulgare L.), and expressed an epitope-tagged version of the cDNA in Arabidopsis, a species with no CSLH genes and no ␤-glucan in its walls. Transgenic Arabidopsis lines that had detectable amounts of the epitopetagged HvCSLH1 protein accumulated ␤-glucan in their walls. The presence of ␤-glucan was confirmed by immunoelectron microscopy (immuno-EM) of sectioned tissues and chemical analysis of wall extracts. In the chemical analysis, characteristic tri-and tetra-saccharides were identified by high-performance anion-exchange chromatography and MALDI-TOF MS following their release from transgenic Arabidopsis walls by a specific ␤-glucan hydrolase. Immuno-EM also was used to show that the epitopetagged HvCSLH1 protein was in the endoplasmic reticulum and Golgi-associated vesicles, but not in the plasma membrane. In barley, HvCSLH1 was expressed at very low levels in leaf, floral tissues, and the developing grain. In leaf, expression was highest in xylem and interfascicular fiber cells that have walls with secondary thickenings containing ␤-glucan. Thus both the CSLH and CSLF families contribute to ␤-glucan synthesis in grasses and probably do so independently of each other, because there is no significant transcriptional correlation between these genes in the barley tissues surveyed.

Plants show varied cellular responses to salinity that are partly associated with maintaining low cytosolic Na+ levels and a high K+/Na+ ratio. Plant metabolites change with elevated Na+, some changes are likely to help restore osmotic balance while others protect Na+-sensitive proteins. Metabolic responses to salt stress are described for two barley (Hordeum vulgare L.) cultivars, Sahara and Clipper, which differed in salinity tolerance under the experimental conditions used. After 3 weeks of salt treatment, Clipper ceased growing whereas Sahara resumed growth similar to the control plants. Compared with Clipper, Sahara had significantly higher leaf Na+ levels and less leaf necrosis, suggesting they are more tolerant to accumulated Na+. Metabolite changes in response to the salt treatment also differed between the two cultivars. Clipper plants had elevated levels of amino acids, including proline and GABA, and the polyamine putrescine, consistent with earlier suggestions that such accumulation may be correlated with slower growth and/or leaf necrosis rather than being an adaptive response to salinity. It is suggested that these metabolites may be an indicator of general cellular damage in plants. By contrast, in the more tolerant Sahara plants, the levels of the hexose phosphates, TCA cycle intermediates, and metabolites involved in cellular protection increased in response to salt. These solutes remain unchanged in the more sensitive Clipper plants. It is proposed that these responses in the more tolerant Sahara are involved in cellular protection in the leaves and are involved in the tolerance of Sahara leaves to high Na+.

The walls deposited by growing pollen tubes contain two types of β-glucan, the (1,3)-β-glucan callose and the (1,4)-β-glucan cellulose, as well as various α-linked pectic polysaccharides. Pollen tubes of Nicotiana alata Link et Otto, an ornamental tobacco, were therefore used to identify genes potentially encoding catalytic subunits of the callose synthase and cellulose synthase enzymes. Reverse transcriptase-polymerase chain reactions (RT-PCR) with pollen-tube RNA and primers designed to conserved regions of bacterial and plant cellulose synthase (CesA) genes amplified a fragment that corresponded to an abundantly expressed cellulose-synthase-like gene named NaCslD1. A fragment from a true CesA gene (NaCesA1) was also amplified, but corresponding cDNAs could not be identified in a pollen-tube library, consistent with the very low level of expression of the NaCesA1 gene. RT-PCR with pollen-tube RNA and primers designed to regions conserved between the fungalFKS genes [that encode (1,3)-β-glucan synthases] and their presumed plant homologs (the Gsl or glucan-synthase-like genes) amplified a fragment that corresponded to an abundantly expressed gene named NaGsl1. A secondGsl gene detected by RT-PCR (NaGsl2) was expressed at low levels in immature floral organs. The structure of full-length cDNAs of NaCslD1, NaCesA1, and NaGsl1 are presented. Both NaCslD1and NaGsl1 are predominantly expressed in the male gametophyte (developing and mature pollen and growing pollen tubes), and we propose that they encode the catalytic subunits of two β-glucan synthases involved in pollen-tube wall synthesis. Different β-glucans deposited in one cell type may therefore be synthesized by enzymes from different gene families.

Self-incompatibility (SI) is a genetic mechanism that restricts inbreeding in flowering plants. In the nightshade family (Solanaceae) SI is controlled by a single multiallelic S locus. Pollen rejection in this system requires the interaction of two S locus products: a stylar (S)-RNase and its pollen counterpart (pollen S). pollen S has not yet been cloned. Our understanding of how this gene functions comes from studies of plants with mutations that affect the pollen but not the stylar SI response (pollen-part mutations). These mutations are frequently associated with duplicated S alleles, but the absence of an obvious additional allele in some plants suggests pollen S can also be deleted. We studied Nicotiana alata plants with an additional S allele and show that duplication causes a pollen-part mutation in several different genetic backgrounds. Inheritance of the duplication was consistent with a competitive interaction model in which any two nonmatching S alleles cause a breakdown of SI when present in the same pollen grain. We also examined plants with presumed deletions of pollen S and found that they instead have duplications that included pollen S but not the S-RNase gene. This finding is consistent with a bipartite structure for the S locus. The absence of pollen S deletions in this study and perhaps other studies suggests that pollen S might be required for pollen viability, possibly because its product acts as an S-RNase inhibitor.S elf-incompatibility (SI) in many plant families is controlled by a multiallelic S locus that enables a style to reject any pollen expressing the same S allelic specificity as itself (1). In the Solanaceae, the family that includes tobacco, tomato, and petunia, SI is described as gametophytic because the allelic specificity of each pollen grain is determined by its own haploid genotype. The S locus in this family encodes a secreted extracellular RNase [stylar (S)-RNase] that accumulates in the style (2). Recognizing which S allele each pollen grain expresses is thought to require an interaction between the S-RNase and an unknown product(s) of a second S locus gene called pollen S (3, 4).As part of a strategy to identify pollen S, we isolated Nicotiana alata plants with gamma ray induced mutations that specifically affect the SI phenotype of pollen but not the SI phenotype of the style (5). Such plants are called pollen-part mutants (PPMs). Because ionizing radiation can cause either the deletion of part of a chromosome or chromosomal aberrations such as translocations, inversions, and fragments (6), the mutations in PPMs are likely to be complex because they can arise through one of a few different types of lesion.Among the PPMs described so far, the most frequent types of lesion are either translocations or small ''centric'' fragments (short extra chromosomes) that carry a duplicated copy of an S allele (5, 7-10). Breakdown of the pollen SI response in these plants occurs because of a ''competitive interaction'' that enables pollen with two different S alleles (but not two i...