ABSTRACT
The line W138 of Petunia hybrida has variegated flowers because it is homozygous for the mutable an1-W138 allele. Excision of the element, causing instability, depends on the presence of the activatorAct1. The previously characterised non-autonomous element dTph1 excises from the dfrC gene in response to Act1. This implies that both non-autonomous elements belong to the same transposable element family. In a range of distantly related cultivars we could detect a single functional Act1 element. Linkage analysis for 11 of these lines showed that Act1 was located on chromosome I in all cases, indicating that the element might be fixed in the genome. A group of cultivars that did not exhibit Act1 activity could be traced back to a recent common origin ('Rose of Heaven'). Cultivars within this group presumably harbour the same inactivated Act1 element. Among the lines tested were 7 lines representing the two species (P. axillaris and P. integrifolia) from which P. hybrida originated. None of these exhibited Act1 activity. We assume that Act1 is present in an inactive state in these lines and that it was activated upon interspecific crossing. In general, lines representing the two parental species and P. hybrida cultivars contain between 5 and 25 dTph1 elements. The lines R27 and W138, however, contain significantly more dTph1 elements (> 50) than all other lines.
ABSTRACT
The fast-moving anionic peroxidase isoenzyme variant PRXa was purified from leaves of petunia (Petunia hybrida). Over 1300-fold purification was achieved by subjecting extracellular extracts to two sequential acetone precipitations and resuspending the pellets at pH 5.0 and pH 8.0, respectively, followed by gel filtration and chromatofocusing. The purified enzyme had an absorbance ratio (A405 nm/A280 nm) of 3.6, a molecular mass of about 37 kDa and a pI of 3.8. Three molecular forms with slightly different molecular masses were separated by concanavalin-A--Sepharose affinity chromatography, indicating that these three forms differ in their carbohydrate moieties. The absorption spectrum of PRXa had maxima at 496 and 636 nm and a Soret band at 405 nm. Spectra of compounds I and IV were obtained by titrating a batch of PRXa stored for several months at -20 degrees C with H2O2. The addition of 1 mol H2O2/mol freshly purified PRXa caused the formation of compound II, indicating that freshly isolated PRXa contains a bound hydrogen donor which is lost upon storage. Compound III was obtained from both preparations in the presence of excess H2O2. The pH optimum of PRXa for the reaction with H2O2 and guaiacol was 5.0 and its specific activity 61 mkat/g protein. Among various aromatic compounds, coniferyl alcohol was polymerized by PRXa to presumed lignin-like material. The extracellular localization and high affinity of PRXa for the cinnamic acid derivatives suggest that this isoenzyme functions in the polymerization or cross-linking of lignin in the plant cell wall.
Subject(s)
Isoenzymes/isolation & purification , Peroxidases/isolation & purification , Plants/enzymology , Catalysis , Chromatography, Affinity , Electrophoresis, Polyacrylamide Gel , Guaiacol/antagonists & inhibitors , Guaiacol/chemistry , Isoelectric Focusing , Isoenzymes/metabolism , Molecular Weight , Peroxidases/metabolism , Peroxides/chemistry , Phenols/metabolism , Spectrum Analysis , Substrate SpecificityABSTRACT
A highly specific rabbit antiserum raised against peroxidase (PRXa) from petunia (Petunia hybrida) was used to investigate the antigenic relatedness of peroxidases in the Solanaceae. After SDS-PAGE of crude leaf extracts from a large number of species of this family, immunoblotting revealed that cross-reacting protein bands were present in all species tested. In order to determine whether these protein bands represent peroxidases, the peroxidase isoenzymes in thorn apple (Datura stramonium L.), tobacco (Nicotiana tabacum L.), sweet pepper (Capsicum annuum L.), potato (Solanum tuberosum L.), and tomato (Lycopersicon esculentum Mill.) were further analyzed. Immunoblots obtained after native PAGE revealed that the antiserum only recognized fast-moving peroxidase isoenzymes that are localized in the apoplast. Despite their serological relatedness, these peroxidases differed with respect to heat stability and apparent molecular weight. Differences in avidity for the petunia PRXa antiserum were suggested by immunoprecipitation with antibodies bound to protein A-Sepharose. The antiserum did not react with peroxidases from horseradish (Armoracea rusticana Gaertn., Mey and Scherb), turnip (Brassica napus L.), African marigold (Tagetes cresta L.), maize (Zea mays L.), and oats (Avena sativa L.). Apparently, the Solanaceae contain orthologous genes encoding the fast-moving anionic peroxidases homologous to petunia PRXa.
ABSTRACT
In Petunia interaction is described between a trans-regulatory genetic element Bi and responsive anthocyanin alleles, resulting in a variegated flower colour pattern. Crosses segregated white-flowering plants that had lost the Bi element and could be shown to contain a responsive allele by the colour pattern resulting from interaction with the standard variegated parent. A weak mutant of the Bi element is described. Some pure lines of Petunia contained the Bi element, others did not. In two lines it could be mapped to chromosome I.
ABSTRACT
The structural gene prxE, coding for a slow cathodic peroxidase in Petunia, has been located to chromosome II, linked to F1. The presence of two mobility alleles in Petunia hybrida can be ascribed to its hybrid descent. Some properties of peroxidase e are mentioned. A gene prxJ is postulated for a still slower cathodic band. The gene Rp1, regulating the onset of expression of the allele prxB2, has been located on chromosome VII (gene order Rp1-prxF-An4). A synopsis of the isoperoxidases and the corresponding genes is given.
ABSTRACT
The structural geneprxD inPetunia codes for a slow moving anodic peroxidase whose activity is sensitive to high concentrations of hydrogen peroxide. The PRXd enzyme could be found in mature and old leaf and stem tissue of full-grown flowering plants. PRXd was found to be absent in tissues from flower corolla and root. The geneprxD is the fourth gene that codes for peroxidases in leaf and stem. Two mobility variants of the PRXd enzyme have been found among our inbred lines using starch gel system II electrophoresis. The geneprxD could be located on chromosome III by a four-point-cross involving the genesprxA, prxD, Mf1 andHt1. The order of the genes established is:Ht1 - Mf1 - prxD - prxA.
ABSTRACT
Two alleles of the structural gene prxA from Petunia, prxA6 and prxA7, could be identified by their differential temporal expression. The alleles prxA6 and prxA7 code for peroxidases with a similar electrophoretic mobility as the products of the previously described alleles prxA1 and prxA5, respectively. The former two alleles differ in that they have a different temporal expression with regard to the temporal expression of the allele prxA2. Crossing experiments indicated that the mutations involved are (cisacting) internal site mutations. In the case of the allele prxA6, the experiments indicated a difference with respect to the allele prxA1 in responsiveness to the action of a trans-acting factor.
ABSTRACT
In root and flower corolla tissue of Petunia several anodic moving peroxidase isoenzymes are present, which cannot be detected in other organs. Alleles of the gene prxF control the presence or absence of several peroxidases that are only present in flower corolla tissue. Alleles of the gene prxG code for two peroxidases that can only be detected in root tissue. In addition to mutations of prxG that cause a change in the electrophoretic mobility of the PRXg enzymes, a mutation was also found that causes the absence of expression in enzyme activity. Crossing experiments indicated that this mutation is located in the gene prxG. Peroxidases encoded by the gene prxH were only found in root tissue. Two alleles of prxH were identified by electrophoretic variation; one allele is responsible for a single band, whereas the other allele could be recognized by a double-banded phenotype. The double-banded PRXh phenotype is suggested to be caused by tandem duplication, followed by mutation in one of the genes. A third prxH allele could be identified by the absence of PRXh activity. The genes prxF, prxG, and prxH were shown to be located on chromosome VII, with the following gene order: prxG-An4-lapB-gpiB-prxH-prxF.
ABSTRACT
Antibodies were raised against the peroxidases encoded by the allele prxA1 to determine the specific activities of the peroxidases encoded by the alleles prxA1, prxA2, prxA3, and prxA5. The results from double diffusion experiments indicated that all peroxidases encoded by the four alleles are antigenically identical. By rocket immuno electrophoresis it was shown that the peroxidases encoded by the alleles prxA1, prxA2, prxA3, and prxA5 have different specific activities. The results presented are discussed in relation to differential expression of the alleles involved.
ABSTRACT
In P. hybrida and the putative progenitor species P. axillaris s. l. and P. integrifolia s. l. five mobility alleles of the structural gene prxA were found. The five alleles show differential expression during development of tissue and plant, caused by internal site mutations. Analysis of young but not yet flowering plants heterozygous for prxA showed that the allele prxA3 is expressed first, followed by the alleles prxA2, prxA5, prxA4 and prxA1, in that order. In mature leaves of young flowering plants the prxA2 allele has the highest expression level, followed by the alleles prxA3, prxA5, prxA4 and prxA1. In mature and old leaves of old plants the expression level of the alleles prxA2, prxA3, prxA4 and prxA5 is about equal, whereas that of the prxA1 allele is about twice as high. Crossing experiments suggested that between the plants used no external site differences exist that cause clearly detectable changes in developmental allozyme balance. Fast moving anodic peroxidases were detected that have a variable mobility over a considerable distance. The probability that these enzymes are precursors of the PRXa enzymes is discussed.
ABSTRACT
In Petunia four alleles of the gene prxB could be identified by starch gel electrophoresis. Investigation of PRXb allozyme balance during development of tissue and plant of prxB heterozygotes showed differential temporal expression of the four mobility alleles. Analysis of developmental allozyme balance of prxB2/B3 heterozygotes from F1 and F2 progenies indicated the presence of a trans-acting regulatory gene termed Rp1. Segregation for the gene Rp1 indicated monogenic inheritance. Independent segregation of the genes Rp1 and prxB was found. When homozygous and recessive, the gene Rp1 speeds up the start of expression in enzyme activity of the prxB2 allele. The alleles prxB1 and prxB4 are expressed later in development than the alleles prxB2 and prxB3, no matter the Rp1 genotype. The final expression level of the prxB alleles is different for all four alleles, irrespective of the Rp1 genotype.
ABSTRACT
By starch gel electrophoresis three mobility variants of a cathodic moving doublet of bands, encoded by the structural gene prxC, were detected in all organs of flowering petunias. In root tissue two of the variants showed a lower electrophoretic mobility than in other organs. During development of flower buds the PRXc enzymes showed an increase in mobility. The gene prxC was located on chromosome IV by showing linkage to the genes An3 and Dw1, by trisomic segregation, and by the construction of triply heterozygous trisomics IV. The gene order on chromosome IV is B1-An3/Dw1-prxC. It was concluded that the temporal programming difference in the expression of the alleles prxC2 and prxC3 is caused by internal site mutation. Analysis of progeny obtained by crossing of lines to the trisomic IV with genotype prxC1/C1/C2 showed differential expression of the two prxC1 alleles of the trisomic IV.
ABSTRACT
Three electrophoretic variants of the peroxidase b isoenzymes in Petunia have been found. The encoding gene prxB is shown to be located on chromosome I by its linkage with the gene Hfl. Analysis of prxB heterozygotes showed a gradual increase of the electrophoretic mobility of all three PRXb allozymes during development and differential expression in enzyme activity of three prxB alleles. The location of prxB on chromosome I was confirmed by an allelic dosage effect in trisomies I, trisomie segregation and the construction of trisomies I with triple-banded PRXb phenotype. From telotrisomic analysis it was concluded that prxB and Hfl are located on the same arm of chromosome I. The unexpected linkage of prxB and Hfl with the gene Fl in one of the crosses was suggested to be caused by a translocation in line SI, involving the gene Fl.
ABSTRACT
As detected by starch gel electrophoresis, the fast moving anodal group of peroxidase isoenzymes, the PRXa complex, of a Petunia homozygous for the encoding gene can be made up of one to four bands, depending on the tissue sampled, the age of the tissue and of the plant, and the genetic background. Additional evidence is presented showing that the PRXa complex is encoded by one structural gene, prxA, rather than by tandem duplicated genes. On the basis of electrophoretic variation in Petunia hybrida and related species, five prxA alleles were found. A prxA internal site mutation was found recognized by the absence of recombination between the mutation that affected the temporal programme of the gene and the mutation that altered the mobility of the enzyme. By a three-point test, the gene prxA was located on chromosome III and found to be linked to the genes Mf1 and Ht1 in the order prxA-Mf1-Ht1. The construction of a trisomic III triply heterozygous for prxA confirmed the location of prxA.
ABSTRACT
A gene termed gpiB, coding for one of the two isoenzyme zones of glucose phosphate isomerase in Petunia, has been mapped to a locus on chromosome VII by means of linkage to the marker An4, and by an allelic dosage effect on enzyme activity in trisomics. The high degree of linkage of electrophoretic alleles of gpiB to the pollen colour allele pair An4/an4, as demonstrated in the ancestral species, P. axillaris s.l. and P. integrifolia s.l., has been conserved in all cultivars of P. hybrida investigated. Another gene, coding for the enzyme leucyl-aminopeptidase could also be mapped to chromosome VII and the gene order An4 - lapB - gpiB determined. Apparently, distribution of lapB alleles is not related to the hybrid descent of P. hybrida.
Subject(s)
Amino Acid Oxidoreductases/genetics , Escherichia coli/genetics , Alanine Dehydrogenase , Chromosome Mapping , Genes , Mutation , Operon , PhenotypeABSTRACT
The peroxidase system in Petunia can be separated into a considerable number of bands by starch gel electrophoresis. The electrophoretic pattern obtained is not only dependent on the genotype of the plant but also on the organ investigated, the state of development of that organ, and the age of the plant. Several structural and regulatory genes seem to be involved in the production of the peroxidase isoenzymes. In Petunia at least three structural genes can be distinguished by their independent segregation. It is suggested that a few more structural genes code for the peroxidase isoenzymes in Petunia. In addition, post-transcriptional modification may contribute to the number of peroxidase isoenzymes.
ABSTRACT
The sequence of three genes involved in cell division in E. coli has been determined to be ftsA-envA-azi by three-point transduction experiments. An ftsA envA double mutant strain forms filaments at the restrictive temperature of 42 degrees C, and not chains, but, like the chain forming envA parent strain, is hypersensitive to rifampicin.
Subject(s)
Escherichia coli , Genes , Mitosis , Chromosome Mapping , Chromosomes, Bacterial , GenotypeABSTRACT
Some properties of a d-glutamic acid auxotroph of Escherichia coli B were studied. The mutant cells lysed in the absence of d-glutamic acid. Murein synthesis was impaired, accompanied by accumulation of uridine-5'-diphosphate-N-acetyl-muramyl-l-alanine (UDP-MurNac-l-Ala), as was shown by incubation of the mutant cells in a cell wall medium containing l-[(14)C]alanine. After incubation of the parental strain in a cell wall medium containing l-[(14)C]glutamic acid, the acid-precipitable radioactivity was lysozyme degradable to a large extent. Radioactive UDP-MurNac-pentapeptide was isolated from the l-[(14)C]glutamic acid-labeled parental cells. After hydrolysis, the label was exclusively present in glutamic acid, the majority of which had the stereo-isomeric d-configuration. Compared to the parent the mutant incorporated less l-[(14)C]glutamic acid from the wall medium into acid-precipitable material. Lysozyme degraded a smaller percentage of the acid-precipitable material of the mutant than of that of the parent. No radioactive uridine nucleotide precursors could be isolated from the mutant under these conditions. Attempts to identify the enzymatic defect in this mutant were not successful. The activity of UDP-MurNac-l-Ala:d-glutamic acid ligase (ADP; EC 6.3.2.9) (d-glutamic acid adding enzyme) is not affected by the mutation. Possible pathways for d-glutamic acid biosynthesis in E. coli B are discussed.
