RESUMEN

A lectin isolated from the roots of the legume, Dolichos biflorus, binds to Nod factors produced by rhizobial strains that nodulate this plant and has a deduced amino acid sequence with no significant homology to any lectin reported to date. This lectin also is an enzyme that catalyzes the hydrolysis of phosphoanhydride bonds of nucleoside di- and triphosphates; the enzyme activity is increased in the presence of carbohydrate ligands. This lectin-nucleotide phosphohydrolase (LNP) has a substrate specificity characteristic of the apyrase category of phosphohydrolases, and its sequence contains four motifs characteristic of this category of enzymes. LNP is present on the surface of the root hairs, and treatment of roots with antiserum to LNP inhibits their ability to undergo root hair deformation and to form nodules on exposure to rhizobia. These properties suggest that this protein may play a role in the rhizobium-legume symbiosis and/or in a related carbohydrate recognition event endogenous to the plant.

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RESUMEN

Seven genomic fragments encoding isoforms of tomato (Lycopersicon esculentum) plasma membrane H(+)-ATPase were cloned and characterized. Genomic DNA gel-blot analysis indicated that probes corresponding to LHA1 through LHA7 hybridized to a common set of seven to nine restriction fragments at moderate stringency and to single, distinct fragments at high stringency. RNA gel-blot and polymerase chain reaction (PCR)-based RNA analyses indicated that LHA1, LHA2, and LHA4 transcripts were present in all organs examined (roots, hypocotyls, stems, immature leaves, mature leaves, green fruit, and red ripe fruit). LHA1 mRNA was present at similar abundance in all organs, LHA2 mRNA was most abundant in hypocotyls and leaves, and LHA4 mRNA was most abundant in roots and hypocotyls. RNA gel-blot and RNA-based PCR assays indicated that LHA3, LHA5, LHA6, and LHA7 mRNA was present at very low or nondetectable levels in all organs, suggesting that these genes are either expressed at very low levels or in organs not examined or that they are regulated by hormonal or environmental cues that were not tested. Indoleacetic acid (IAA) treatment of tomato hypocotyl segments resulted in modest changes in abundance of LHA1, LHA2, and LHA4 transcripts, but these changes were not correlated with the time course of IAA-induced growth. In addition, constitutively silent LHA genes were not activated by IAA. These results indicate that at least seven genomic sequences are present in tomato that may encode plasma membrane H(+)-ATPases, at least three of which are expressed relatively abundantly at the mRNA level.

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Calcium-dependent regulatory mechanisms participate in diverse developmentally, hormonally, and environmentally regulated processes, with the precise control of cytosolic Ca2+ concentration being critical to such mechanisms. In plant cells, P-type Ca(2+)-ATPases localized in the plasma membrane and the endoplasmic reticulum are thought to play a central role in regulating cytoplasmic Ca2+ concentrations. Ca(2+)-ATPase activity has been identified in isolated plant cell membranes, but the protein has not been characterized at the molecular level. We have isolated a partial-length cDNA (LCA1) and a complete genomic clone (gLCA13) encoding a putative endoplasmic reticulum-localized Ca(2+)-ATPase in tomato. The deduced amino acid sequence specifies a protein (Lycopersicon Ca(2+)-ATPase) of 1048 amino acids with a molecular mass of 116 kDa, eight probable transmembrane domains, and all of the highly conserved functional domains common to P-type cation-translocating ATPases. In addition, the protein shares approximately 50% amino acid sequence identify with animal sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPases but less than 30% identity with other P-type ATPases. Genomic DNA blot hybridization analysis indicates that the Lycopersicon Ca(2+)-ATPase is encoded by a single gene. RNA blot hybridization analysis indicates the presence of three transcript sizes in root tissue and a single, much less abundant, transcript in leaves. Lycopersicon Ca(2+)-ATPase mRNA levels increase dramatically upon a 1-day exposure to 50 mM NaCl. Thus this report describes the primary structure of a higher-plant Ca(2+)-ATPase and the regulation of its mRNA abundance by salt stress.

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RESUMEN

Two cDNA clones (LHA1 and LHA2) from tomato (Lycopersicon esculentum) which likely encode isoforms of the plasma membrane H(+)-ATPase were isolated. The longest cDNA (3229 base pairs), LHA1, comprises an open reading frame that encodes a 956 amino acid, 105 kilodalton polypeptide with several potential transmembrane domains. In vitro transcription and translation of LHA1 yields a major translation product of approximately 100 kilodaltons that is immunoprecipitable with antiserum to the corn root plasma membrane H(+)-ATPase. LHA2 encodes a portion of a coding sequence that is 96% identical to LHA1, suggesting that LHA2 encodes an isoform of the H(+)-ATPase. Genomic DNA gel blot analysis indicates that both LHA1 and LHA2 hybridize to a common set of six to eight restriction fragments at moderate stringency and to single distinct fragments at high stringency. LHA1 and LHA2 map to distinct sites on chromosomes three and six, respectively. RNA gel blot analysis indicates that both LHA1 and LHA2 hybridize to 3.4 kilobase pair transcripts present in both leaves and roots, although the LHA2 transcript is relatively more abundant in leaves than in roots. These results indicate that in tomato as many as six to eight genes may encode the plasma membrane H(+)-ATPase, two of which are expressed at the level of mRNA in both roots and leaves.