Expression of the developmentally regulated catalase (cat) genes in maize

The catalase (H₂O₂:H₂O₂ oxidoreductase; E.C.1.11.1.6; CAT) gene-enzyme system in Zea mays L (maize) represents an ideal model for studying the molecular basis of developmental gene regulation in higher eukaryotes. This system comprises a family of structural genes that are highly regulated, both temporally and spatially, during maize development. In maize, there are four distinct forms (isozymes) of catalase that are readily discernible by convetional separation procedures. Three of the catalases have been studied in detail from a genetic and biochemical viewpoint. The catalases CAT-1, CAT-2, and CAT-3 are encoded by the distinct, unlinked genes Cat1, Cat2, and Cat3, respectively. Each of the structural genes is highly regulated both spatially and temporally in its expression. Cat1 is expressed primarily in the endosperm, aleurone, pericarp, and scutellum of developing kernels, and in the root, shoot, and scutellum of very young seedlings. Cat2 is expressed primarily in the scutellum and leaf during postgerminative sporophytic development. Cat3 is expressed, for the most part, in the shoot and pericarp of young seedlings. A number of regulatory variants have been recovered that affect the developmental program of expression of the catalases. Analysis of one variant allowed for the identification of a temporal regulatory gene (Car1) that specifically alters the developmental program of the Cat2 structural gene by acting to regulate the rate of CAT-2 protein synthesis. Cat1 has been mapped on chromosome 1S, 37 map units (m.u.) from the Cat2 structural gene. Another variant line has been isolated which lacks expression of the Cat2 gene in its tissues at all stages of development. Isolated polysomes from this line (A16) were translated in vitro, and the products were immunoprecipitated with CAT-2-specific antibodies. No CAT-2 was detectable in the A16 labeled immunoprecipitates, whereas CAT-2 was readily detected in the normal line, W64A, under similar conditions. The temporal and spatial expression of the Cat structural genes is not only influenced by genetic factors (as above), but is also responsive to exogenously applied environmental signals: light, hormones, and temperature. The mechanisms by which such signals specifically affect CAT-2 expression will be discussed.     

Antioxidant Defences under Hyperoxygenic and Hyperosmotic Conditions in Leaves of Two Lines of Maize with Differential Sensitivity to Drought

The effects of hyperoxygenic and hyperosmotic stress on severalaspects of antioxidant defences were studied in the leaves ofa drought-sensitive (LG11) and a drought-tolerant (LIZA) lineof maize (Zea mays L.). When leaf disks were subjected to theseverest stress conditions (100% O₂ and 0.5 M mannitol), theactivities of antioxidant enzymes, such as superoxide dismutase(SOD), catalase (Cat), ascorbate peroxidase (Asc-Px) and glutathionereductase (GSSG-Red), remained higher in disks of LIZA thanin disks of LG11. The ratios of activities of SOD to Cat, SODto Asc-Px and SOD to GSSG-Red were much higher in leaf disksfrom LG11 than in those from LIZA. Damage, as indicated by,for example, the extend of lipid peroxidation, the destructionof chlorophyll and carotenoids, the decrease in levels of proteinsulfhydryl groups and the leakage of electrolytes from cellswas apparent in leaf disks of both LIZA and LG11 as consequenceof the applied stresses. However, the damage was less markedin LIZA than in LG11. (Received September 2, 1992; Accepted July 20, 1993)     

Effect of Defoliation During the Early Vegetative Phase and at Silking on Growth of Maize (Zea mays L.)

Two experiments were conducted in the glasshouse and in thefield to evaluate the effect of leaf loss on development, drymatter accumulation and yield of maize. In the glasshouse, defoliationtreatments were imposed on maize after 3 weeks of planting.Removing 2 or 3 leaves every 2 weeks affected plant height,days to tassel, root and shoot yield. The plant diameter androot:shoot ratio were not affected by defoliation. In the field experiment, six defoliation treatments were imposed7 days after 50 per cent silking. Defoliation of all leaveswas the most severe treatment on cob d. wt, dry matter accumulationin grains, weight of 100 test grains and yield. The effect ofremoving all leaves above the ear was not significantly differentfrom that of removing all leaves below the ear. The effect ofremoving half of the leaves above the ear was not differentfrom the control. Zea mays, vegetative growth, dry matter accumulation, yield, defoliation     

Biological control of southern corn leaf blight by Trichoderma atroviride SG3403

Trichoderma atroviride SG3403 showed high biocontrol activity against southern corn leaf blight (SCLB; pathogen: Cochliobolus heterostrophus). T. atroviride SG3403 could cause death of C. heterostrophus race O hypha on plates. Spraying T. atroviride SG3403 conidia suspension over maize seedling leaves protected the corn from SCLB infection. Biocontrol effect lasted for 30 days in the field. Trichoderma strain was able to induce resistance response in corn leaves against pathogen infection. In corn leaves treated with T. atroviride SG3403, the enzyme activities of phenylalanine ammonia lyase (PAL) and superoxide dismutase (SOD) reached the highest at 24 h, enzyme activity of catalase (CAT) reached the highest at 36 h after inoculation of pathogen C. heterostrophus race O. RNA expression levels of Pal, Sod and Cat (which synthesis enzyme PAL, SOD and CAT) were also upregulated and corresponded to the enzyme activity at the same time point. Enzyme activities and corresponding genes expression induced by Trichoderma SG3403 was more obvious than that induced by pathogen only, which implies that T. atroviride SG3403 induced corn defense gene expression against pathogen infection. Thus, induced resistance mechanism was possibly involved in the biocontrol of SCLB by T. atroviride SG3403.     

Analysis of variants affecting the catalase developmental program in maize scutellum

Summary The catalase of maize scutella is coded for by two loci, Cat1 and Cat2, which are differentially expressed in this tissue during early seedling growth. Two variant lines have been previously identified in which the developmental program for the expression of the Cat2 structural gene in the scutellum has been altered. Line R6–67 exhibits higher than normal levels of CAT-2 catalase in this tissue after four days of postgerminative growth. This phenotype is controlled by a temporal regulatory gene designated Car1. Line A16 exhibits a CAT-2 null phenotype. Further analysis of Car1 verifies the initial indication that it is trans-acting and exhibits strict tissue (scutellum) specificity. A screen of other available inbred lines uncovered eight additional catalase high-activity lines. All eight lines exhibit significantly higher than normal levels of CAT-2 protein. Two of these lines have been shown to be regulated by Car1 as in R6–67. Another line (A338) uncovered during the screen exhibits a null phenotype for CAT-2 protein and resembles A16. Catalase activity levels are low in the scutellum and no CAT-2 CRM (cross-reacting material) is present in the tissues of this line. Also, unlike most maize lines, CAT-2 cannot be induced in the leaf tissue of A338 upon exposure to light. Finally, a single line (A337), demonstrating a novel catalase developmental program, was identified.     

Differential expression of the maize catalase genes during kernel development: the role of steady-state mRNA levels

In maize three isozymic forms of catalase, CAT-1, CAT-2, and CAT-3 are encoded by three distinct and unliked structural genes (Cat1, Cat2 and Cat3). Catalase activity profiles and zymogram analysis were used to examine the spatial and temporal expression of the three genes during kernel maturation. Three developmental stages of catalase expression were observed in the growing kernel. During stage 1 (6-12 days after pollination), both Cat1 and Cat3 were expressed; during stage 2 (15-18 days after pollination) only Cat1 expression was observed; and during stage 3 (21-30 days after pollination), Cat1 and Cat2 were expressed. The major constituent tissues of the kernel were examined to determine their contribution to total kernel catalase expression. Each of the tissues was found to have a unique pattern of catalase gene expression. RNA blot analysis, using catalase gene-specific nucleic acid probes, suggests that the differential expression of the three catalase genes observed in the kernel is regulated by controlling the distribution of steady-state mRNA species for the three genes.     

Developmental Analysis of Steady-State Levels of Cu/Zn and Mn Superoxide Dismutase mRNAs in Maize Tissues

The superoxide dismutase (Sod) steady-state mRNA levels in maizeseedlings and developing kernels were examined by RNA blot analysis,using Cu/Zn and Mn cDNA probes encoding the cytosolic and mitochondrialSOD isozymes, respectively. The cytosolic (Cu/Zn) Sod steady-statemRNA levels remained relatively constant for the various tissuesexamined. In contrast, the mRNA levels of the mitochondrial(Mn) SOD-3 isozyme increased in postgerminative scutella. Thesteady-state mRNA of the Atp2 gene, F₁ ATPase ß subunit,was compared to the Sod3 (Mn) mRNA levels. Results of this comparativestudy suggest that the steady-state levels of mRNAs transcribedby nuclear genes encoding essential mitochondrial proteins areindependently regulated throughout development. (Received April 2, 1990; Accepted September 10, 1990)     

Pre-flowering Growth and Development of the Inflorescences of Maize: II. ACCUMULATION AND PARTITIONING OF DRY MATTER AND NITROGEN BY INFLORESCENCES

The accumulation and partitioning of dry matter and nitrogenwere examined in the developing tassel and two uppermost earshoots of field-grown maize under varying levels of appliednitrogen and times of sowing. Accumulation of dry matter and nitrogen within an axillary branchalways favoured the ear over the husk and shank. Dry matterand nitrogen accumulated faster in the first ear than in thetassel or second ear and the partitioning between inflorescencesof dry matter and nitrogen was not affected by treatment. Therelative rate of growth, RGR(dry matter), of the first and secondear shoots increased by up to 42% at high levels of appliednitrogen and with early sowing. In contrast, the relative rateof accumulation of nitrogen (RNAR) was not sensitive to N supply,although it was reduced, on average, by 22% at the late timeof sowing. We conclude that accumulation, but not partitioning, of drymatter and nitrogen between developing inflorescences of maizeare altered by nitrogen application, time of sowing, and positionof the inflorescence on the stem. Key words: Maize, N-application, partitioning, inflorescence, sowing time     

Molecular evolution of maize catalases and their relationship to other eukaryotic and prokaryotic catalases

We have compared the nucleotide and protein sequences of the three maize catalase genes with other plant catalases to reconstruct the evolutionary relationship among these catalases. These sequences were also compared with other eukaryotic and prokaryotic catalases. Phylogenies based on distances and parsimony analysis show that all plant catalases derive from a common ancestral catalase gene and can be divided into three distinct groups. The first, and major, group includes maizeCatl, barleyCat1, riceCatB and most of the dicot catalases. The second group is an apparent dicot-specific catalase group encompassing the tobaccoCat2 and tomatoCat. The third is a monocot-specific catalase class including the maize Cat3, barley Cat2, and riceCatA. The maize Cat2 gene is loosely related to the first group. The distinctive features of monocot-specific catalases are their extreme high codon bias at the third position and low degree of sequence similarity to other plant catalases. Similarities in the intron positions for several plant catalase genes support the conclusion of derivation from a common ancestral gene. The similar intron position between bean catalases and human catalase implies that the animal and plant catalases might have derived from a common progenitor gene sequence. Correspondence to: J.G. Scandalios     

Analysis of Organ Specificity of a Low Temperature Responsive Gene Family in Rye (Secale cereale L.)

The barley (Hordeum vulgare L.) low temperature responsive geneblt14 was used as a probe, to isolate two different cognateclones (rlt1412; rlt1421) from a rye (Secale cereale L.) cDNAlibrary prepared from low temperature-treated (6°C day/2°C night) shoot meristems of the cultivar, Puma. Northernblot analysis revealed that low temperature expression of rlt1412is highest in root tissues whereas, rlt1421 shows greatest mRNAaccumulation in mature leaf tissues. There is a relationshipbetween the steady-state levels of these mRNA species and thefrost hardiness of Puma (North American cultivar) and Rhayader(UK cultivar) such that the expression ofboth genes is higherin the more frost hardy cultivar, Puma, compared with Rhayader. DNA and predicted amino acid sequence analysis indicated thatthe rye and barley clones encode small proteins with consensusN-terminal signal sequences whose biological function is atpresent unknown. The relevant sequences are lodged in the EMBL data base. Key words: Rye, cold, cDNA, organ specificity, low temperature genes     

Catalase Activity and Post-anoxic Injury in Monocotyledonous Species

Three anoxia-intolerant species, Glyceria maxima, Juncus effususand Iris germanica (var. Quechei), and three anoxia-tolerantspecies Schoenoplectus lacustris, Acorus calamus and Iris pseudacoruswere chosen for investigation. Rhizomes of anoxia-intolerantspecies show increased catalase activities when returned toair after periods of prolonged anoxia. Levels of catalase remainedfairly constant in anoxia-tolerant species under the same conditions.In the anoxia intolerant G. maxima, the post-anoxic increasein catalase activity was reduced by circulating the anaerobicatmosphere. This treatment also reduced the ethanol contentof the tissue under incubation, and increased the survival ofthe rhizomes as seen in their ability to resume growth in thepost-anoxic phase. Exposure of anaerobic G. maxima rhizomesto ethanol vapour increased post-anoxic levels of catalase activityand when this produced a 5-fold increase always resulted indeath of the rhizomes. Acetaldehyde vapour applied in the sameway gave rise to increases in catalase activity followed byrapid death of the rhizomes. It is suggested that post-anoxic oxidation of anaerobicallyaccumulated ethanol may result in a surge of acetaldehyde production,which could exert a toxic effect on the recovering tissues.The possible role of catalase in an ethanol-oxidation reaction,which is well documented in animals, is discussed in the lightof the association between the natural accumulation of largeconcentrations of ethanol and subsequent post-anoxic death insome plant tissues. Key words: Catalase, post-anoxia, ethanol     

Molecular cloning,characterization and expression analysis of two catalase isozyme genes in barley

Clones representing two distinct barley catalase genes, Cat1 and Cat2, were found in a cDNA library prepared from seedling polysomal mRNA. Both clones were sequenced, and their deduced amino acid sequences were found to have high homology with maize and rice catalase genes. Cat1 had a 91% deduced amino acid sequence identity to CAT-1 of maize and 92% to CAT B of rice. Cat2 had 72 and 79% amino acid sequence identities to maize CAT-2 and-3 and 89% to CAT A of rice. Barley, maize or rice isozymes could be divided into two distinct groups by amino acid homologies, with one group homologous to the mitochondria-associated CAT-3 of maize and the other homologous to the maize peroxisomal/glyoxysomal CAT-1. Both barley CATs contained possible peroxisomal targeting signals, but neither had favorable mitochondrial targeting sequences. Cat1 mRNA occurred in whole endosperms (aleurones plus starchy endosperm), in isolated aleurones and in developing seeds, but Cat2 mRNA was virtually absent. Both mRNAs displayed different developmental expression patterns in scutella of germinating seeds. Cat2 mRNA predominated in etiolated seedling shoots and leaf blades. Barley genomic DNA contained two genes for Cat1 and one gene for Cat2. The Cat2 gene was mapped to the long arm of chromosome 4, 2.9 cM in telomeric orientation from the mlo locus conferring resistance to the powdery mildew fungus (Erysiphe graminis f.sp. hordei).