The carotenoid and abscisic acid content of viviparous kernels and seedlings ofZea mays L.

Carotenoid and abscisic acid (ABA) levels were determined in endosperm, embryos and seedlings of wild-type and viviparous (vp) mutants ofZea mays L. Carotenoid concentrations were determined by absorption spectrometry following purification by high-performance liquid chromatography and ABA concentrations by combined gas chromatography-mass spectrometry. Lutein and zeaxanthin were the terminal carotenoids in wild-type tissue. The carotenoid profiles ofvp-1 andvp-8 tissue were similar to that of the wild type; invp-2, vp-5, vp-7 andvp-9 carotenogenesis was blocked at early stages so that xanthophylls were absent. Except forvp-1, where the ABA content was similar to the wild type, the ABA content ofvp embryos was substantially reduced, to 6–16% of the corresponding wild type. Thus, the absence of xanthophylls was associated with reduced ABA content, which was in turn correlated with vivipary. Kernels ofvp-8 had a reduced ABA content although xanthophylls were present. Seedlings of carotenoid-deficient mutants rescued from viviparous kernels contained less ABA than did wild-type seedlings grown in the same way. Furthermore, the ABA concentration of such seedlings did not increase in response to water deficit. Conversely,vp-1 seedlings contained normal levels of carotenoids and ABA. Carotenoid-deficient seedlings did not contain appreciable amounts of chlorophyll so that chloroplast development was not normal. Thus ABA-deficiency could be associated with abnormal plastid development rather than the absence of carotenoids per se.Abbreviations ABA abscisic acid - DAP days after pollination - i.d. internal diameter - FW fresh weight - GC-MS combined gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - MS mass spectrometry - vp viviparous     

Internode length in Zea mays L.

[¹³C, ³H]Gibberellin A₂₀ (GA₂₀) has been fed to seedlings of normal (tall) and dwarf-5 and dwarf-1 mutants of maize (Zea mays L.). The metabolites from these feeds were identified by combined gas chromatography-mass spectrometry. [¹³C, ³H]Gibberellin A₂₀ was metabolized to [¹³C, ³H]GA₂₉-catabolite and [¹³C, ³H]GA₁ by the normal, and to [¹³C, ³H]GA₂₉ and [¹³C, ³H]GA₁ by the dwarf-5 mutant. In the dwarf-1 mutant, [¹³C, ³H]GA₂₀ was metabolized to [¹³C, ³H]GA₂₉ and [¹³C, ³H]GA₂₉-catabolite; no evidence was found for the metabolism of [¹³C, ³H]GA₂₀ to [¹³C, ³H]GA₁. [¹³C, ³H]Gibberellin A₈ was not found in any of the feeds. In all feeds no dilution of ¹³C in recovered [¹³C, ³H]GA₂₀ was observed. Also in the dwarf-5 mutant, the [¹³C]label in the metabolites was apparently undiluted by endogenous [¹³C]GAs. However, dilution of the [¹³C]label in metabolites from [¹³C, ³H]GA₂₀ was observed in normal and dwarf-1 seedlings. The results from the feeding studies provide evidence that the dwarf-1 mutation of maize blocks the conversion of GA₂₀ to GA₁.Abbreviations GA_n gibberellin A_n - GC-MS combined gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - RP reverse phase     

Graviresponsiveness and abscisic-acid content of roots of carotenoid-deficient mutants of Zea mays L.

The abscisic-acid (ABA) content of roots of the carotenoid-deficient w-3, vp-5, and vp-7 mutants of Z. mays was analyzed using gas chromatography-mass spectrometry with an analysis sensitivity of 6 ng ABA g^–1 fresh weight (FW). Roots of normal seedlings of the same lines were characterized by the following amounts of ABA (as ng ABA g^–1 FW,±standard deviation): w-3, 279±43; vp-5, 237±26; vp-7, 338±61. We did not detect any ABA in roots of any of the mutants. Thus, the lack of carotenoids in these mutants correlated positively with the apparent absence of ABA. Primary roots of normal and mutant seedlings were positively gravitropic, with no significant differences in the curvatures of roots of normal as compared with mutant seedlings. These results indicate that ABA 1) is synthesized in maize roots via the carotenoid pathway, and 2) is not necesary for positive gravitropism by primary roots of Z. mays.Abbreviation ABA abscisic acid     

The phytochrome system in light-grown Zea mays L.

The phytochrome system is analyzed in light-grown maize (Zea mays L.) plants, which were prevented from greening by application of the herbicide SAN 9789. The dark kinetics of phytochrome are not different in the first, second or third leaf. It is concluded that in light-grown maize plants phytochrome levels are regulated by P_r formation and P_fr and P_r destruction, rather than by P_frP_r dark reversion. P_r undergoes destruction after it has been cycled through P_fr. The consequences of this P_r destruction on the phytochrome system are discussed.Abbreviations SAN 9789 4-chloro-5-(methylamino)-2-(,,-trifluoro-m-tolyl)-3(2H) pyridazinone - P_fr far-red absorbing form of phytochrome - P_r red absorbing form of phytochrome - P_tot P_fr+P_r     

Leaf vasculature in Zea mays L.

The vascular system of the Zea mays L. leaf consists of longitudinal strands interconnected by transverse bundles. In any given transverse section the longitudinal strands may be divided into three types of bundle according to size and structure: small, intermediate, large. Virtually all of the longitudinal strands intergrade structurally however, from one bundle type to another as they descend the leaf. For example, all of the strands having large-bundle anatomy appear distally as small bundles, which intergrade into intermediates and then large bundles as they descend the leaf. Only the large bundles and the intermediates that arise midway between them extend basipetally into the sheath and stem. Most of the remaining longitudinal strands of the blade do not enter the sheath but fuse with other strands above and in the region of the blade joint. Despite the marked decrease in number of longitudinal bundles at the base of the blade, both the total and mean cross-sectional areas of sieve tubes and tracheary elements increase as the bundles continuing into the sheath increase in size. Linear relationships exist between leaf width and total bundle number, and between cross-sectional area of vascular bundles and both total and mean cross-sectional areas of sieve tubes and tracheary elements.     

A diazotrophic bacterial endophyte isolated from stems of Zea mays L. and Zea luxurians Iltis and Doebley

The apoplastic fluids of field-grown Zea mays and Zea luxurians plants were isolated from surface sterilized stem tissue by centrifugation and spread on agar plates containing a nitrogen-free, defined medium. The predominant bacterium isolated from these plates was characterized further. The ability of this bacterium to fix nitrogen was confirmed by its ability to grow on a semi-solid, nitrogen-free medium and reduce ¹⁵N₂ to ¹⁵NH₃ and acetylene to ethylene. Protions of the nifH and 16S rRNA genes from this organism were amplified by PCR and sequenced. The nifH gene, which codes for dinitrogenase reductase, from this organism is closely related to nifH from Klebsiella pneumoniae. Similarly, the 16S rRNA gene sequences and carbon utilization tests grouped it closely with K. pneumoniae. Based an these data, the isolates from Z. mays and Z. luxurians are tentatively classified as Klebsiella spp. (Zea). The ability of this bacterium to contribute to the nitrogen economy of the corn plant is unknown.     

Effects of salinity and benzyl adenine on development and function of microhairs of Zea mays L

Bicellular microhairs are present on the surfaces of leaves of grasses with the exception of the Pooideae. In some halophytic grasses, these glandular hairs secrete salt, suggesting the intriguing question ‘can the microhairs of grasses that do not normally encounter salinity also secrete salt?’ Microhairs were counted in replicas of the adaxial and abaxial surfaces of leaves of various ages of maize plants growing either in the absence of salt or in the presence of 40, 80 or 120 mM NaCl. The number of microhairs per unit area of adaxial leaf surface of the youngest leaf almost doubled as the salinity increased from zero to 120 mM NaCl; on the abaxial surface, the number of microhairs increased by 50%. Spraying this leaf with benzyl adenine (BA) caused, when averaged across salinities and surfaces, a 32% increase in the number of microhairs. Salinity reduced leaf area but in all the salinity treatments, spraying with BA increased the total number of microhairs per leaf. Washing leaves of plants provided estimates of the loss of salt from those leaves. There were large differences between the Na:K molar ratios in the washing solution and the leaf tissue, indicating a high selectivity for sodium over potassium for loss from the leaf. BA did not influence the efficiency of salt loss, expressed per microhair, at any salinity level, but did increase loss per leaf. Thus, BA increased salt loss from plants due to its influence on the number of microhairs and leaf area, but not due to its effect on the efficiency of the secretion process per se.     

Tubular extensions of the plasmalemma in leaf cells of Zea mays L.

Leaf tissues of Zea mays were examined with a transmission electron microscope and a high-voltage electron microscope. Tubular extensions (invaginations) of the plasmalemma were found in vascular parenchyma cells and thick-walled, lateformed sieve elements of intermediate and small veins, and in epidermal, mesophyll, and sheath cells of all leaves examined. No continuity seems to exist between the tubules and other cellular membranes.     

Photosynthetic carbon metabolism during leaf ontogeny in Zea mays L.: Enzyme studies

The activities of several enzymes, including ribulose-1,5-diphosphate (RuDP) carboxylase (EC 4.1.1.39) and phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) were measured as a function of leaf age in Z. mays. Mature leaf tissue had a RuDP-carboxylase activity of 296.7 mol CO₂ g^-1 fresh weight h^-1 and a PEP-carboxylase activity of 660.6 mol CO₂ g^-1 fresh weight h^-1. In young corn leaves the activity of the two enzymes was 11 and 29%, respectively, of the mature leaves. In senescent leaf tissue, RuDP carboxylase activity declined more rapidly than that of any of the other enzymes assayed. On a relative basis the activities of NADP malic enzyme (EC 1.1.1.40), aspartate (EC 2.6.1.1) and alanine aminotransferase (EC 2.6.1.2), and NAD malate dehydrogenase (EC 1.1.1.37) exceeded those of both PEP and RuDP carboxylase in young and senescent leaf tissue. Pulse-chase labeling experiments with mature and senescent leaf tissue show that the predominant C₄ acid differs between the two leaf ages. Labeling of alanine in senescent tissue never exceeded 4% of the total ¹⁴C remaining during the chase period, while in mature leaf tissue alanine accounted for 20% of the total after 60 s in ¹²CO₂. The activity of RuDP carboxylase during leaf ontogeny in Z. mays parallels the development of the activity of this enzyme in C₃ plants.Abbreviations RuDP ribulose-1,5-diphosphate - PEP phosphoenol pyruvate - PGA 3-phosphoglycerate     

Intercellular compartmentation of sucrose synthesis in leaves of Zea mays L.

The incorporation of ¹⁴C into sucrose and hexose phosphates during steady-state photosynthesis was examined in intact leaves of Zea mays L. plants. The compartmentation of sucrose synthesis between the bundle sheath and mesophyll cells was determined by the rapid fractionation of the mesophyll and comparison of the labelled sucrose in this compartment with that in a complete leaf after homogenisation. From these experiments it was concluded that the majority of sucrose synthesis occurred in the mesophyll cell type (almost 100% when the time-course of sucrose synthesis was extrapolated to the time of ¹⁴C-pulsing). The distribution of enzymes involved in sucrose synthesis between the two cell types indicated that sucrose-phosphate synthetase was predominantly located in the mesophyll, as was cytosolic (neutral) fructose-1,6-bisphosphatase activity. Stromal (alkaline) fructose-1,6-bisphosphatase activity was found almost exclusively in the bundle-sheath cells. No starch was found in the mesophyll tissue. These data indicate that in Zea mays starch and sucrose synthesis are spatially, separated with sucrose synthesis occurring in the mesophyll compartment and starch synthesis in the bundle sheath.     

The intercellular compartmentation of metabolites in leaves of Zea mays L.

Sap extracted from attached leaves of two-to three-week-old maize plants witt the aid of a roller device was almost devoid of bundle-sheath contamination as judged by the distribution of mesophyll and bundle-sheath markers. The extraction could be done very rapidly (less than 1 s) and the extract immediately quenched in HClO₄ or reserved for enzyme assay. Comparison of the contents of metabolites in intact leaves and in the leaf extract allowed estimation of the distribution of metabolites between the bundle-sheath and the mesophyll compartments. Substantial amounts of metabolites such as malate and amino acids were present in the non-photosynthetic cells of the midrib. In the illuminated leaf, triose phosphate was predominantly located outside the bundle-sheath while the major part of the 3-phosphoglycerate was in the bundle sheath. The results indicate the existence of concentration gradients of triose phosphate and 3-phosphoglycerate in the leaf which are capable of maintaining carbon flow between the mesophyll and bundle-sheath cells during photosynthesis. There was no evidence for the existence of a gradient of pyruvate between the bundle-sheath and the mesophyll cells.     

Regulation of flower development in cultured ears of maize (Zea mays L.)

Summary Young ears of maize were cultured in two different liquid media containing either kinetin (KN) or kinetin + gibberellic acid (KN + GA₃) in order to manipulate stamen and gynoecium development. In KN medium, stamens developed and gynoecia aborted in the flowers of the cultured immature ears. In the KN + GA₃ medium, however, ovaries with silks developed and stamens aborted. These differential morphological events were recorded with SEM photomicrographs at regular intervals after excision of ear inflorescences. In addition, the mitotic activity in the developing or aborting organs was determined over a 75-h period. It increased from 6% to 14% in developing organs (i.e. stamens in KN medium, and gynoecia in KN + GA₃ medium) and gradually decreased to 1% in the degenerating organs (i.e. gynoecia in KN medium, and stamens in KN + GA₃ medium) by 45 h of culture. The mitotic activity reached zero in degenerating flower organs by 75 h of culture. Whether these differential sensitivities to the exogenously applied members of these two plant growth regulator classes are unique to our in vitro system or reflect a more general control feature of in vivo inflorescences must await further clarification.     

Carbon metabolism and gas exchange in leaves of Zea mays L.

The aim of this work was to investigate the mechanism of formation of triose phosphates and 3-phosphoglycerate during photosynthetic induction in leaves of Zea mays. Simultaneous measurements of gas exchange, chlorophyll a fluorescence and metabolite contents of maize leaves were made. Leaves illuminated in the absence of CO₂ showed a build-up of triose phosphates during the first 2 min of illumination which was comparable to the build-up observed in the presence of CO₂. Isolated mesophyll protoplasts, which lack the Calvin cycle, also showed a build-up of triose phosphates upon illumination. Leaves contained amounts of phosphoglycerate mutase and enolase adequate to account for the formation of triose phosphates and 3-phosphoglycerate from intermediates of the C₄ cycle and their precursors.     

Lateral hydraulic conductivity of early metaxylem vessels in Zea mays L. roots

The hydraulic conductivity of the lateral walls of early metaxylem vessels (Lp_x in m · s^–1 · MPa^–1) was measured in young, excised roots of maize using a root pressure probe. Values for this parameter were determined by comparing the root hydraulic conductivities before and after steam-ringing a short zone on each root. Killing of living tissue virtually canceled its hydraulic resistance. There were no suberin lamellae present in the endodermis of the roots used. The value of Lp_x ranged between 3 · 10^–7 and 35 · 10^–7 m · s^–1 · MPa^–1 and was larger than the hydraulic conductivity of the untreated root (Lp_r = 0.7 · 10^–7 to 4.0 · 10^–7 m · s^–1 · MPa^–1) by factor of 3 to 13. Assuming that all flow through the vessel walls was through the pit membranes, which occupied 14% of the total wall area, an upper limit of the hydraulic conductivity of this structure could be given(Lp_pm=21 · 10^–7 to 250 · 10^–7 m · s^–1 · MPa^–1). The specific hydraulic conductivity (Lp_cw) of the wall material of the pit membranes (again an upper limit) ranged from 0.3 · 10^–12 to 3.8 · 10^–12 m² · s^–1 · MPa^–1 and was lower than estimates given in the literature for plant cell walls. From the data, we conclude that the majority of the radial resistance to water movement in the root is contributed by living tissue. However, although the lateral walls of the vessels do not limit the rate of water flow in the intact system, they constitute 8–31% of the total resistance, a value which should not be ignored in a detailed analysis of water flow through roots.Abbreviatations and Symbols k_wr (T ₁ ^2/W ) rate constant (half-time) of water exchange across root (s^–1 or s, respectively) - Lp_cw specific hydraulic conductivity of wall material (m² · s^–1 · MPa^–1) - Lp_pm hydraulic conductivity of pit membranes (m · s ^–1 · MPa^–1) - Lp_r hydraulic conductivity of root (m · s^–1 · MPa^–1) - Lp_x lateralhydraulic conductivity of walls of root xylem (m · s ^–1 · MPa^–1) This research was supported by a grant from the Bilateral Exchange Program funded jointly by the Natural Sciences and Engineering Research Council of Canada and the Deutsche Forschungsgemeinschaft to C.A.P., and by a grant from the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 137, to E.S. The expert technical help of Mr. Burkhard Stumpf and the work of Ms. Martina Murrmann and Ms. Hilde Zimmermann in digitizing chart-recorder strips is gratefully acknowledged.     

Differential expression of a hydroxyproline-rich cell-wall protein gene in embryonic tissues of Zea mays L.

A hydroxyproline-rich glycoprotein (HRGP) component of the maize cell wall was shown to be present in different organs of the plant by extraction of cell wall proteins and detection by Western blotting and immunocytochemistry. Antibodies raised against the protein or against synthetic peptides designed from the protein sequence immunoprecipitated a proline-rich polypeptide which was synthesized in-vitro from poly(A) ⁺ RNA extracted from different tissues of the plant and from the complete in-vitro-transcribed mRNA. A very low amount of the protein was found in immature embryos. In particular, the protein could not be detected in the scutellum either by Western blotting or by immunocytochemistry. In agreement with this finding, HRGP mRNA was barely detected in the scutellum, in contrast to its accumulation in the embryo axis. Our results indicate the existence of a unique cell wall structure in embryonic tissues from maize as well as a tissuespecific component of the control of maize HRGP gene expression, distinct to others already described such as cell division.Abbreviations HRGP(s) hydroxyproline-richglycoprotein(s) - DAP days after pollination The present work was supported by grants from Plan Nacional de Investigation Cientifica y Técnica (grant BI088-0242) and European Communities (grant BAP-374). L.R.-A. is the recipient of a fellowship from the Plan Nacional de Formación de Personal Investigador.     

Distribution and structure of the plasmodesmata in mesophyll and bundle-sheath cells of Zea mays L.

In leaf blades of Zea mays L. plasmodesmata between mesophyll cells are aggregated in numerous thickened portions of the walls. The plasmodesmata are unbranched and all are characterized by the presence of electron-dense structures, called sphincters by us, near both ends of the plasmodesmatal canal. The sphincters surround the desmotubule and occlude the cytoplasmic annulus where they occur. Plasmodesmata between mesophyll and bundle-sheath cells are aggregated in primary pit-fields and are constricted by a wide suberin lamella on the sheath-cell side of the wall. Each plasmodesma contains a sphincter on the mesophyll-cell side of the wall. The outer tangential and radial walls of the sheath cells exhibit a continuous suberin lamella. However, on the inner tangential wall only the sites of plasmodesmatal aggregates are consistently suberized. Apparently the movement of photosynthetic intermediates between mesophyll and sheath cells is restricted largely or entirely to the plasmodesmata (symplastic pathway) and transpirational water movement to the cell walls (apoplastic pathway).Abbreviation ER endoplasmic reticulum     

Early responses to gibberellic acid in a dwarf maize mutant (Zea mays L. d 1)

Early effects of gibberellic acid (GA₃) (1–4 h treatment) on the ion ratios in a dwarf maize mutant (Zea mays L. d ₁) showing normal growth after hormone treatment, have been investigated by electron microprobe analysis. GA₃ exerts a different effect on the ion ratios in plastids, cytoplasm and vacuoles in short term experiments. The Cl content of chloroplasts and cytoplasm increases without a lag phase after GA₃ treatment. The K content of plastids increases after a lag phase of 2 h, whereas in the cytoplasm an increase can be observed immediately after GA₃ addition. The hormone has only little influence on the Ca content of the cell compartments investigated. Control experiments with water and the physiologically inactive GA₃ methylester confirm the specifity of the short-term actions of GA₃ on the ion ratios. The primary action of GA₃ at the membrane level is discussed.     

Growth,graviresponsiveness and abscisic-acid content of Zea mays seedlings treated with Fluridone

Ten-d-old seedlings of Zea mays L. cv. Tx 5855 treated with 1-methyl-3-phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1H)-pyridinone (Fluridone) were analyzed for abscisic acid (ABA) content using high-performance liquid chromatography with an analysis sensitivity of 2.5 ng ABA g^-1 fresh weight (FW). Seedlings were divided into three portions: leaves, detipped roots, and root tips (terminal 1.5 mm). Control plants (water treatment only; no Fluridone) were characterized by the following amounts of ABA: leaves, 0.114±0.024 (standard deviation) g ABA g^-1 FW; detipped roots, 0.260±0.039±g ABA g^-1 FW; root tips, no ABA detected. We did not detect any ABA in tissues of Fluridone-treated plants. Primary roots of treated and untreated seedlings were strongly graviresponsive, with no significant differences between the curvatures or the growth rates of primary roots of Fluridone-treated and control seedlings. These results indicate that 1) Fluridone completely inhibits ABA synthesis, and 2) ABA is not necessary for positive gravitropism by primary roots of Zea mays.Abbreviations ABA abscisic acid - Fluridone 1-methyl-3-phenyl-5-(3-[trifluoromethyl]phenyl)-4-(1H)-pyridinone - FW fresh weight - SD standard deviation