The Plant Journal

Jackson, David; Roberts, Keith; Martin, Cathie

doi: 10.1111/j.1365-313X.1992.00425.xpmid: N/A

The co‐ordination of expression of anthocyanin biosynthetic genes was studied in developing flowers. Four genes encoding enzymes operating late in the anthocyanin biosynthetic pathway are induced together during flower development but the early steps appear to be induced more rapidly. Co‐ordination of expression could imply a common regulatory mechanism controlling the expression of metabolically related genes. The data presented here show that while four genes may share such a mechanism for the control of their expression during flower development, different control processes regulate the early steps of the pathway. Spatially, gene expression is patterned across the flower and appears to be very similar for all the biosynthetic genes. However, the observed influence of the regulatory gene Delila shows that the spatial co‐ordination of gene expression must involve more than one regulatory system. Delila itself appears to have a dual function, being required for activation of expression of the later genes in the flower tube but repressing chalcone synthase gene expression in the mesophyll of the corolla lobes. It is postulated that common signals induce the expression of genes in the pathway during flower development. The data presented here suggest that the same regulatory mechanism interprets these signals for four of the genes encoding the later biosynthetic enzymes, but that different or modified mechanisms interpret the signals to control expression of chalcone synthase and chalcone isomerase genes in Antirrhinum flowers.

Nambara, Eiji; Naito, Satoshi; McCourt, Peter

doi: 10.1111/j.1365-313X.1992.00435.xpmid: N/A

In order to investigate the role of the plant hormones gibberellin (GA) and abscisic acid (ABA) in seed development and germination the GA biosynthetic inhibitor, Uniconazol, was used to isolate mutants with abnormal germination profiles. In one of these mutants, the ability to germinate on Uniconazol is due to a mutation in the ABI3 gene. However, unlike the previously reported abi3 mutant, this line displays an array of seed‐specific developmental defects. The accumulation of seed reserve proteins is dramatically reduced due to reduced levels of the storage protein mRNA. The embryos remain green throughout development and are desiccation intolerant. However, immature seeds are completely non‐dormant and grow normally. These results suggest the ABI3 gene is essential for the synthesis of seed storage proteins and for the protection of the embryo during desiccation.

D'Amico, Leone; Valsasina, Barbara; Daminati, M. Gloria; Fabbrini, M. Serena; Nitti, Gianpaolo; Bollini, Roberto; Ceriotti, Aldo; Vitale, Alessandro

doi: 10.1111/j.1365-313X.1992.00443.xpmid: 1344885

Treatment of developing bean cotyledons with the inhibitor of N ‐glycosylation tunicamycin enhanced the synthesis of at least two polypeptides with molecular mass 78 kDa and 97 kDa. Pulse‐chase experiments and subcellular fractionation indicated that these are endoplasmic reticulum (ER) residents. The 78 kDa protein is a major component of the ER protein fraction and, by N‐terminal sequencing, was identified as a bean homolog of the mammalian 78 kDa glucose‐regulated protein (GRP78). This is a molecular chaperone that is probably involved in the folding and oligomerization of several animal and yeast proteins in the ER. When newly synthesized storage glyco‐proteins phaseolin, phytohemagglutinin or α‐amylase inhibitor were immunoprecipitated from an ER preparation of tunicamycin‐treated tissue, the GRP78 homolog was always co‐precipitated. Bound GRP78 homolog could be released by ATP treatment. These results suggest that, at least when glycosylation is inhibited, this protein plays a role in the early stages of the synthesis of vacuolar storage proteins.

Kanevski, Ivan F.; Thakur, Sanjay; Cosowsky, Laurey; Sunter, Garry; Brough, Clare; Bisaro, David; Maliga, Pal

doi: 10.1111/j.1365-313X.1992.00457.xpmid: N/A

The feasibility of obtaining clonal lines with replicating, multicopy geminivirus vectors by direct DNA trans‐formation of cultured tobacco cells was studied. The replicating vectors pTGA32 and pST31 are based on the tomato golden mosaic virus (TGMV) A genome and encode the neomycin phosphotransferase type II (NPT‐II) enzyme that confers kanamycin resistance to plant cells. Following introduction into plant cells, unit‐length viral genomes were released from the tandem repeats and replicated. In protoplasts, replication of unit‐length pTGA32 and pST31 was about as efficient as replication of unit‐length DNA A from plasmid pTGA26, which contains 1.5 copies of wild‐type DNA A. Tobacco suspension culture cells were bombarded with the recombinant DNA A constructs and selected for kanamycin resistance. The number of kanamycin‐resistant clones per bombardment was about the same when the TGMV DNA A vectors or a non‐replicating plasmid (pLC14) which also encodes NPT‐II was used. Replicating, unit‐length DNA A in up to approximately 1000 copies per cell was found in about 10% of the kanamycin‐resistant clones selected following bombardment of cells with TGMV vectors. The results suggest that geminiviruses may serve as useful multicopy vectors in cultured cells.

Pröls, F.; Meyer, P.

doi: 10.1046/j.1365-313X.1992.t01-20-00999.xpmid: 1344886

The regions of integration of a transferred DNA‐fragment from three transgenic Petunia hybrida plants were analysed for their influence on the expression of the foreign DNA. Each of the three transformants, lines 16, 17 and 24, contained a fragment of a plasmid on which two genes were located, an npt‐II gene which renders the plants resistant to kanamycin and the A1 gene from Zea mays, a visible marker gene that leads to the production of a brick red anthocyanin pigment in the flowers. Inactivation of both genes in line 16 is associated with integration into a region of highly repetitive DNA, while the integration sites of the other two lines were essentially unique. The integration regions of lines 17 and 24, both of which show expression of the foreign genes at characteristically different intensities, showed a distinct methylation pattern that was stably conserved for these regions in both trangenic and wild‐type plants. The characteristic methylation pattern of the two integration regions was also imposed on the border region of the integrated fragments and might thus be responsible for the differences in the intensity of gene expression observed among the two lines.

Lorberth, Ruth; Dammann, Christian; Ebneth, Marcus; Amati, Simone; Sánchez‐Serrano, Jose J.

doi: 10.1046/j.1365-313X.1992.t01-21-00999.xpmid: N/A

The proteinase inhibitor II (pin2) gene family exhibits two different modes of expression. It is, on the one hand, constitutively expressed in flowers of potato and tomato plants, and in potato tubers. On the other hand, its expression is induced in the plant foliage by mechanical wounding. To define cis‐regulatory elements involved in pin2 promoter activity, deletion analysis of a potato pin2 promoter has been performed in stably and transiently transformed potato and tobacco plants. Two different elements, a quantitative enhancer and a regulatory element, are required for promoter activity. While functional promoter elements required for pin2 activity in tubers and wounded leaves could not be separated, its expression in flowers is mediated by different cis‐acting sequences. Induction of pin2 expression in leaves by treatment with the plant growth regulators abscisic acid and jasmonic acid, and the general metabolite sucrose, depends on the presence of the regulatory element involved in expression in tubers and wounded leaves. Thus, pin2 expression in tubers and wounded leaves apparently results from the action of similar hormonal signals on closely linked promoter elements, while a different signal pathway leads to its constitutive expression in flowers.

Handberg, Kurt; Stougaard, Jens

doi: 10.1111/j.1365-313X.1992.00487.xpmid: N/A

In the Leguminosae plant family, few of the individual plant species have been used for plant molecular biology research. Among the species investigated no obvious representative ‘model’ legume has emerged. Here a member of the tribe Loteae, Lotus japonicus (Regel) Larsen is proposed as a candidate. L. japonicus is a diploid, autogamous species, with a good seed set, and a generation time of approximately 3 months. The haploid genome consists of six chromosomes and the genome size was estimated to be relatively small (0.5 pg per haploid complement). L. japonicus is susceptible to Agrobacterium tumefaciens and transgenic plants can be regenerated after hygromycin or kanamycin selection. Tissue culture conditions and procedures for transformation and regeneration are described. Stable transformation is demonstrated by segregation of the hygromycin selectable marker after selfing of transgenic plants or test crosses. The possibility of mapping polymorphic DNA markers inbred lines of L. japonicus is also discussed.

Nasrallah, Mikhail E.; Kandasamy, Muthugapatti K.; Nasrallah, June B.

doi: 10.1111/j.1365-313X.1992.00497.xpmid: N/A

A new locus, SCF1, which regulates S‐locus function has been identified in Brassica campestris. The spontaneous scf1 mutation described in this paper causes the stigma to be receptive to all pollen including selfpollen, but does not affect the pollination phenotype of the male gametophyte. In the stigmas of scf1 homozygotes, the S‐locus‐linked SLG gene and two other unlinked members of the S‐multigene family, all of which encode secreted glycoproteins, are coordinately down‐regulated at the RNA level, while the expression of the S‐locus‐derived SRK gene, which encodes a putative receptor protein kinase, is unaffected. It is proposed that the SCF1 locus encodes a positive trans‐acting factor required for the enhanced transcription of several genes in the stigmatic papillar cells during pistil maturation. The effects of the scf1 mutation provide strong support for the involvement of the cloned S‐genes in the operation of the pollen‐stigma interaction of incompatibility in B. campestris and related crucifer species.

Jansen, Marcel A.K.; Sessa, Guido; Malkin, Shmuel; Fluhr, Robert

doi: 10.1111/j.1365-313X.1992.00507.xpmid: N/A

Styles nurture and guide pollen tubes to the ovules. The styles of Nicotiana tabacum, a C3 plant, contain a concentric strand of transmitting tract cells replete with well‐developed chloroplasts. It is shown that the chloroplasts have normal ultrastructure and electron transport ability. However, they were found to be devoid of Rubisco, the key enzyme responsible for carbon fixation in C3 plants. Nevertheless, non‐invasive fluorescence techniques showed a light‐driven photosynthetic flux. Carbon fixation via phosphoenol pyruvate carboxylase (PEPC) into malate was demonstrated, the latter accumulating during stylar development. Characterization of stylar PEPC in vitro and in vivo revealed apparent Km values consistent with bicarbonate as a rate limiting factor for photosynthetic flux. Presumably, in the closed confines of the intact style, respired CO2 is the source of carbonate. Enhanced photosynthetic flux was detected following pollination, suggesting utilization of the additional respired bicarbonate and underlining metabolic interactions between the style and the elongating pollen tube.

Karrer, Erik E.; Rodriguez, Raymond L.

doi: 10.1046/j.1365-313X.1992.t01-22-00999.xpmid: N/A

Isolated rice embryos were used to investigate the regulatory effects of endosperm extracts and pure sugars on the expression of α‐amylase gene RAmy3D and a sucrose synthase gene homologous to the maize isozyme Ss2. The high‐level expression of RAmy3D in the scutella of isolated embryos could be inhibited by a variety of sugars as well as endosperm extracts from germinated rice grains. Glucose, at a concentration of 250 mM, was most effective in repressing RAmy3D mRNA accumulation. Furthermore, this repression was reversible. Interestingly, RAmy3D repression was always accompanied by the induction of sucrose synthase gene expression. These results support a model in which the expression of α‐amylase and sucrose synthase genes in the rice scutellum are counter‐regulated by the influx of sugars from the endosperm.