A protein involved in co-ordinated regulation of inorganic carbon and glucose metabolism in the facultative photoautotrophic cyanobacteriumSynechocystis PCC6803

The involvement of a gene ofSynechocystis PCC6803,icfG, in the co-ordinated regulation of inorganic carbon and glucose metabolism, was established. TheicfG gene codes for a 72 kDa protein, which shows no homology with those registered in data libraries. Expression oficfG required glucose, the actual inducer probably being glucose-6-phosphate, and was independent of light and of the external inorganic carbon concentration. Mutants carrying an inactivated copy oficfG were constructed. Their growth characteristics were identical to those of the wild type under all regimes except in limiting inorganic carbon with glucose being present either before or after the transfer to the limiting conditions. These conditions completely prevented growth, both in the light and in the dark. The inhibition could be relieved by several intermediates of the tricarboxylic acid cycle. Assays of various enzymic activities related to inorganic carbon uptake and to its assimilationvia either the Calvin cycle or phosphoenolpyruvate carboxylase did not reveal the level of action of IcfG. Possible models include a blockage of the assimilation of both carbon sources in the absence of IcfG, or the inhibition of Ci incorporation route(s) essential under limiting inorganic carbon conditions, even when glucose is present, and even in the dark.     

The S11 and S13 self incompatibility alleles in Solanum chacoense Bitt. are remarkably similar

A genomic clone of the S11 allele from the self-incompatibility locus (S locus) in Solanum chacoense Bitt. has been isolated by cross-hybridization to the S. chacoense S13 allele and sequenced. The sequence of the S11 allele contains all the features expected for S genes of the Solanaceae, and S11 expression, as assessed by northern blots and RNA-PCR, was similar to that of other S. chacoense S alleles. The S11 protein sequence shares 95% identity with the phenotypically distinct S13 protein of S. chacoense and is the gametophytic S allele with the highest similarity to an existing allele so far discovered. Only 10 amino acid changes differentiate the mature proteins from these two alleles, which sets a new lower limit to the number of changes that can produce an altered S allele specificity. The amino acid substitutions are not clustered, suggesting that an accumulation of random point mutations can generate S allele diversity. The S11 intron is unusual in that it could be translated in frame with the coding sequence, thus suggesting an additional mechanism for the generation of new S alleles.     

Meristem-specific gene expression directed by the promoter of the S-phase-specific gene,cyc07, in transgenic Arabidopsis

A genomic clone for the cyc07 gene, which is expressed specifically at the S phase during the cell cycle in synchronous cultures of periwinkle (Catharanthus roseus) cells, was isolated. Determination of the nucleotide sequence of the clone revealed that the cyc07 gene consists of seven exons separated by six introns. Genomic Southern analysis indicated that the cyc07 gene is present as a single copy per haploid genome in periwinkle. Expression of related genes was detected in a wide range of other plants. Transgenic Arabidopsis plants were generated that expressed the gene for -glucuronidase (GUS) under the control of the promoter of the cyc07 gene. The tissue-specific pattern of expression directed by the promoter was investigated by analysis of GUS activity. Histochemical tests demonstrated that 589 bp of the 5-upstream sequence of the cyc07 gene could direct specifical expression of the GUS reporter gene in meristematic tissues in transgenic plants. The spatial pattern of expression directed by the promoter was closely correlated with meristematic activity and cell proliferation, suggesting an association between the function of the cyc07 gene and cell proliferation.     

The use of a hybrid genetic system to study the functional relationship between prokaryotic and plant multi-enzyme fatty acid synthetase complexes

Fatty acid synthesis in bacteria and plants is catalysed by a multi-enzyme fatty acid synthetase complex (FAS II) which consists of separate monofunctional polypeptides. Here we present a comparative molecular genetic and biochemical study of the enoyl-ACP reductase FAS components of plant and bacterial origin. The putative bacterial enoyl-ACP reductase gene (envM) was identified on the basis of amino acid sequence similarities with the recently cloned plant enoyl-ACP reductase. Subsequently, it was unambiguously demonstrated by overexpression studies that theenvM gene encodes the bacterial enoyl-ACP reductase. An anti-bacterial agent called diazaborine was shown to be a specific inhibitor of the bacterial enoyl-ACP reductase, whereas the plant enzyme was insensitive to this synthetic antibiotic. The close functional relationship between the plant and bacterial enoyl-ACP reductases was inferred from genetic complementation of anenvM mutant ofEscherichia coli. Ultimately,envM gene-replacement studies, facilitated by the use of diazaborine, demonstrated for the first time that a single component of the plant FAS system can functionally replace its counterpart within the bacterial multienzyme complex. Finally, lipid analysis of recombinantE. coli strains with the hybrid FAS system unexpectedly revealed that enoyl-ACP reductase catalyses a rate-limiting step in the elongation of unsaturated fatty acids.     

Non-systemic expression of a stress-responsive maize polyubiquitin gene (Ubi-1) in transgenic rice plants

We have used the promoter, 1st exon and 1st intron of the maize polyubiquitin gene (Ubi-1) for rice transformation experiments and revealed the characteristic expression of Ubi-1 gene: (1) Ubi-1 gene is not regulated systemically but rather individual cells respond independently to the heat or physical stress; (2) Ubi-1 gene changes its tissue-specific expression in response to stress treatment; (3) the expression of Ubi-1 gene is dependent on cell cycle.     

The biosynthesis of rosmarinic acid in suspension cultures of Coleus blumei

Suspension cultures of Coleus blumei accumulate very high amounts of rosmarinic acid, an ester of caffeic acid and 3,4-dihydroxyphenyllactate, in medium with elevated sucrose concentrations. Since the synthesis of this high level of rosmarinic acid occurs in only five days of the culture period, the activities of the enzymes involved in the biosynthesis are very high. Therefore all the enzymes necessary for the formation of rosmarinic acid from the precursors phenylalanine and tyrosine could be isolated from cell cultures of Coleus blumei: phenylalanine ammonia-lyase, cinnamic acid 4-hydroxylase, hydroxycinnamoyl:CoA ligase, tyrosine aminotransferase, hydroxyphenylpyruvate reductase, rosmarinic acid synthase and two microsomal 3- and 3-hydroxylases. The main characteristics of these enzymes of the proposed biosynthetic pathway of rosmarinic acid will be described.Abbreviations DHPL 3,4-dihydroxyphenyllactate - DHPP 3,4-dihydroxyphenylpyruvate - pHPL 4-hydroxyphenyllactate - pHPP 4-hydroxyphenylpyruvate - RA rosmarinic acid     

Influence of exogenous abscisic acid on germination and plantlet conversion frequencies of hybrid larch somatic embryos (Larix × leptoeuropaea)

The effect of exogenous abscisic acid, provided to somatic embryos during the maturation step, on endogenous abscisic acid and its main conjugated form (abscisic acid glucose ester), germination and conversion frequencies is presented in this paper. Abscisic acid measurements were obtained after a methanolic extraction, a fractionation through high performance liquid chromatography, quantitation with an immunoassay and identification of the quantitated compound using gas chromatography-mass spectrometry. Results show that endogenous abscisic acid and abscisic acid glucose ester levels are clearly correlated with the exogenous abscisic acid concentration provided to the embryos. Maturation was clearly enhanced by exogenous abscisic acid, but no correlation was found between abscisic acid concentration and germination frequency. Conversely, development of the aerial part of the germinated somatic embryos was dependent upon the abscisic acid concentration in the culture medium and results suggest that this dependence could be related to the endogenous abscisic acid content.