Persistence of differences between peroxidase isoenzymes of flax genotrophs in tissue culture

Anionic peroxidase isoenzymes from seedling root, hypocotyl and cotyledon regions of the large (L) and small (S) flax genotrophs were separated on acrylamide gels. Tissue cultures were initiated from each of these regions of the seedlings, and maintained for a 200-day period with six transfers. The differences in electrophoretic mobility of the peroxidase isoenzymes between L and S noted in seedlings, and also in main stems of adult plants, were still present in the tissue cultures.     

Indoleacetic acid oxidase activity in main stem homogenates of flax genotrophs and genotypes

The IAA-oxidase and peroxidase capabilities along the length of the main stem tissues of two flax genotrophs L and S and two flax genotypes R and M were examined in vitro. Stem gradients for peroxidase activity increased basipetally in all plant types, as did IAA-oxidase activity gradients at non-rate-limiting concentrations of Mn²⁺. Correlations between peroxidase activity and non-rate-limited IAA-oxidase activity supported the contention of dual activities on the same molecule. At rate-limiting concentrations of Mn²⁺, IAA-oxidase activity did not correlate with peroxidase activity. Plant type differences were detected in rate-limited IAA-oxidase activity. This activity was higher in the stem region immediately above the cotyledons (axillary buds) of the more branched types, L and R, than in the sparsely branched types, S and M.     

Relative mobility and activity of leaf malate dehydrogenase in flax(Linum usitatissimum) genotrophs and genotypes

Malate dehydrogenase (MDH) band relative mobility (R _m) and activity were examined in leaf extracts of Durrant's flax genotrophs, L and S, and flax genotypes, R and M. MDH activity in leaves from just below the inflorescence was higher in the two smaller, sparsely branched plant types, S and M, than in the larger, more branched plant types, L and R. The MDH electrophoretic banding pattern in flax leaf extracts consisted of three major anionic bands, MDH-1, MDH-2, and MDH-3. NoR _m differences were detected between corresponding isozymes of genotypes R and M. For the genotrophs, however, all three bands of S migrated faster than the corresponding bands of L. Codominance was absent in F₁ hybrids; SR _m was dominant for MDH-2 and MDH-3 and LR _m was dominant for MDH-1. The observations suggest that MDHR _m in L and S may be controlled by a modifier locus (or loci). Previous studies indicate that a modifier locus may also control heritable genotrophic differences in peroxidase (PER) and acid phosphates (AP)R _m. The three enzyme systems are compared.The financial assistance of the Natural Sciences and Engineering Research Council of Canada is acknowledged with thanks.     

Purification,characterization and catalytic activity of anionic zucchini peroxidase

The isolation and purification, by preparative electrofocusing, of the major anionic (ZPOA) and cationic (ZPOC) isoenzymes, collected from young zucchini squash, are reported. The M _r and sugar content are similar to those found previously for the major isoenzymes from the ripe fruits and in the range commonly observed for plant peroxidases. The amount of the two cationic enzymes was very low compared with that of anionic ZPOA. The anionic enzyme has been characterized by electronic, circular dichroism, proton NMR and electron paramagnetic resonance spectroscopy. The spectra are qualitatively similar to those of the corresponding anionic horseradish peroxidase (HRPA) derivatives, with minor differences attributable to the particular protein environment around the heme. The kinetics of the enzymatic oxidation of a series of phenols by H₂O₂ have been studied. ZPOA shows a parallel behavior to HRPA, but it is systematically more active than HRPA, indicating that the zucchini enzymes have a marked tendency to carry out oxidation of this type of compounds.     

Direct response of peroxidase isozyme relative mobility to short-term environmental temperature changes in flax (Linum usitatissimum)

Abstract Flax plants were transferred from 30°C to 15°C temperatures, and then back to 30°C during their vegetative growth phase. Reciprocal transfers were also made from 15°C to 30°C, and then back to 15°C for an identical period at the same time. Plants were also retained in a constant 30°C or 15°C in growth chambers with identical lighting conditions. The reciprocal transfer period lasted 6 d. The relative mobility (R_m) of an anionic leaf peroxidase isozyme, reflecting its molecular weight/conformation and/or net charge, was monitored over a period starting before the reciprocal transfers and finishing after the return to original temperatures. Two of the four flax types which were studied showed isozyme relative mobility responses related either to the different constant temperatures, or to the transfer from 15°C to 30°C. The responses were not reversible on return to the original lower temperature. Thus, exposure to 30°C, even for a transient 6 d period, produced a one way R_m shift. This suggests a developmental switch-over in the particular form of peroxidase glycoprotein being synthesized, rather than an adaptive response triggered by external environmental changes.     

Possible posttranslational modification,and its genetic control,in flax genotroph isozymes

Environmentally induced flax genotrophs L and S show heritable shifts in the relative mobilities of peroxidase, esterase, and acid phosphatase isozymes, plus a number of nonspecific glycoproteins. All L isozymes migrated faster than corresponding S isozymes in 10% acrylamide gels. Various aspects of these shifts are reviewed here; it is proposed that posttranslational modification, probably of the carbohydrate moieties of these glycoproteins, underlies the shifts. This proposal is discussed in relation to the switch model for genotroph induction.The financial assistance of the Natural Sciences and Engineering Research Council of Canada is acknowledged with thanks.     

Systematic implications of the peroxidase isoenzymes in the Xanthium strumarium complexes

Starch-slab gel electrophoresis showed two patterns of peroxidase isoenzymes in the polymorphic taxon. Xanthium strumarium L. The “strumarium” morphological complex (X. strumarium, in the sense of Millspaugh and Sherff), the putative indigenous plants of the Old World, contained a different pattern from that shown by putative introductions from the New World. Experimental F₁ hybrids between “strumarium” and other complexes, “italicum-pennsylvanicum”, “chinense”, “oviforme” and “cavanillesii”, had the peroxidase pattern of the American plants. These peroxidase isoenzyme patterns were not influenced by the environmental growing conditions, but, along with partial genetic incompatibility, support the taxonomic separation of X. strumarium from other taxa of section Euxanthium.     

Antigenic relationships between petunia peroxidase a and specific peroxidase isoenzymes in other Solanaceae

Summary 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.     

Linkage studies of the h gene with plant weight in flax genotrophs

Summary The association of the H-h (hairy-hairless septa) character with plant weight was studied in the coupling and repulsion phases in F₂ of reciprocal crosses between large (L) and small (S) genotrophs of flax variety Stormont Cirrus. F₂ plants of reciprocal crosses in coupling (L^H x S^h) and in repulsion (L^h x S^H) giving H-h segregations were grown with their parents at two sowing times. Significant positive and negative associations between h and plant weight were obtained. A model is proposed based on the hypothesis that the H phenotype had changed to the h phenotype at the time of induction by a heterochromatic region extending over this locus. In the heterozygote, stable equilibria of the homozygotes are destroyed and transfer of heterochromatin, or number of reiterated sequences, or a decrease in one homologue and an increase in the other, occur in this region between homologous chromosomes. The amount and direction of the association is dependent upon the frequency of transfer: 0% transfer gives complete positive association; 50% transfer, no association; 100% transfer, complete negative association. This mechanism or heterochromatic transfer preserves the Mendelian ratio of 31 of Hh in the F₂. It is also supposed that there must be other controlling elements present as well.     

Kinetics of fenugreek peroxidase isoenzymes

Peroxidase from fenugreek seedlings was separated into 6 isoenzymes; 4 on CM-cellulose and 2 on DEAE-cellulose. The kinetics of these peroxidase isoenzymes with regard to o-dianisidine and catechol are described.     

Partial characterization of peroxidase isoenzymes from rust-affected wheat leaves

Four anodic peroxidase isoenzymes from wheat leaves were purified by column chromatography and their kinetic behavior with common substrates were examined. One isoenzyme is more active in wheat resistant to stem rust fungi and differed from the others in carbohydrate content and also by a specific activity 2–4-fold higher with non-physiological electron donors. As a substrate, eugenol exhibited kinetic behavior different from p-phenylenediamine, guaiacol or o-dianisidine with all isoenzymes. All four isoenzymes showed similar pH and temperature optima and kinetic behavior and apparent K_m values for both H₂O₂ and non-physiological electron donors.     

Changes in peroxidase isoenzymes of Phaseolus mungo hypocotyl cuttings during rooting

A time course study of changes in the pattern of peroxidase isoenzymes shows that two new isoenzymes a and b appeared in hypocotyl cuttings in     

Effect of gamma irradiation on peroxidase isoenzymes during suberization of wounded potato tubers

A comparison of peroxidase isoenzymes in skin, cortex and pith tissues of the potato tuber by thin-layer isoelectric focusing in Sephadex revealed major differences in the isoenzyme patterns. Wounding induced several-fold increases in the peroxidase activity which were correlated with the increased amounts of specific isoenzymes. The anodic and cathodic forms with high activity, normally present in large amounts in skin, were found to be preferentially synthesized in suberizing tissues, suggesting a functional role for peroxidase in the suberization process. Cycloheximide treatment prevented the rapid increase in the content and activity of these specific isoenzymes, which indicated that the increase in peroxidase is due to a de novo synthesis of the enzyme. Suberization is not inhibited by gamma irradiation at sprout-inhibiting dose levels.     

Molecular weight and net charge of peroxidase isozymes in F1 hybrids between L and S flax genotrophs

In the F₁ hybrids between Durrant's L and Durrant's S flax genotrophs, the relative mobilities of the anionic peroxidase isozymes were essentially the same as those in the L parent. The isozymes in both parents and their F₁'s were compared over a range of acrylamide gel concentrations, with plots of log relative mobility against gel concentration. Plots of comparative mobility, relative to the internal standard hemoglobin, against concentration were also examined. Both approaches provided evidence that apparent molecular weight modifications underlay the shift in mobility between the parents and the resemblance of the F₁'s to L, the parent which was homozygous for the dominant alleles controlling the mobility shift for at least two of the isozymes.     

Genetics of the peroxidase isoenzymes in Petunia

Summary 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.     

Cytoplasmic effects on activity of individual anionic isoenzymes of peroxidase in crosses between two genotypes of flax (Linum usitatissimum L.)

Four anionic peroxidase isoenzymes in main stem tissues of flax (Linum usitatissimum L.) were separated electrophoretically on acrylamide gels and their individual activities measured spectrophotometrically in the gels. Activities were expressed in terms of areas of optical density peaks corresponding to the locations of the isoenzymes. Four isoenzymes were assayed for activity in this way in two inbred genotypes and their F₁, F₂, and first backcross progeny. Crosses were made in all combinations between the two parental genotypes and their reciprocal F₁ hybrids to produce 16 progenies of the generations above. Isoenzyme separations and assays for activity were carried out on individual plants of each of the 16 progenies. A model estimating cytoplasmic effects across segregating generations as differences between all progeny of the one compared to the other F₁ reciprocal hybrid, either in a male or a female direction, was fitted to the activity data by weighted least-squares procedures. Cytoplasmic effects transmitted through male gametes from the F₁ reciprocals were demonstrable for three of the four anionic peroxidase isoenzymes.Supported in part by the National Research Council of Canada.