Ptychographic amplitude and phase reconstruction of bichromatic vortex beams.

We experimentally demonstrate that ptychographic coherent diffractive imaging can be used to simultaneously characterize the amplitude and phase of bichromatic orbital angular momenta-shaped vortex beams, which consist of a fundamental field, together with its copropagating second-harmonic field. In contrast to most other orbital angular momentum characterization methods, this approach solves for the complex field of a hyperspectral beam. This technique can also be used to characterize other phase-structured illumination beams, and, in the future, will be able to be extended to other complex fields in the extreme ultraviolet or X-ray spectral regions, as well as to matter waves.

Age-dependent changes in lipid peroxide levels in peripheral organs, but not in brain, in senescence-accelerated mice.

The tissue concentration of lipid peroxides was determined in the brain, heart, liver, lung and kidney of accelerated senescence-prone (SAMP-8) and -resistant (SAMR-1) mice at 3, 6 and 9 months of age by a method involving chemical derivatization and high performance liquid chromatography. The level of lipid peroxides in the brain did not show an age-dependent change, but at each age the brain level of lipid peroxides was significantly higher in SAMP-8 than in SAMR-1. In contrast, the lipid peroxide levels in the peripheral organs showed increases with aging in both strains, and they were significantly higher in SAMP-8 than in SAMR-1 at both 3 and 6 months of age (except at 3 months of age in the kidney). These results suggest that increased oxidative stress in the brain and peripheral organs is a cause of the senescence-related degeneration and impairments seen in SAMP-8.

Cytosolic beta-cyanoalanine synthase activity attributed to cysteine synthases in cocklebur seeds. Purification and characterization of cytosolic cysteine synthases.

The activity of beta-cyanoalanine synthase (CAS, EC 4.4.1.9) in cotyledons of cocklebur seeds (Xanthium pennsylvanicum Wallr.) was detected both in the soluble and particulate fractions. The CAS activity of the soluble fraction (cytosolic CAS activity) was 10 times higher than that of the particulate fraction. The CAS activity of the particulate fraction was confirmed to be localized in the mitochondria. Both enzymatic activities were clearly separated by non-denaturing PAGE. The enzyme with cytosolic CAS activity has been extensively purified and separated into three different forms designated as cyt-1, cyt-2, and cyt-3. According to the SDS-PAGE analysis, the three enzymes are estimated to be a homodimer composed of 35-kDa subunits. The purified enzymes showed CS activity. Partial amino acid sequences of cyt-1 were determined and had a high homology with cysteine synthases (CS, EC 4.2.99.8) from other plant sources. The catalytic action of the purified CSs in converting cyanide and cysteine into H2S and beta-cyanoalanine was confirmed by the detection of significant 14CN incorporation into beta-cyanoalanine. These results indicated that cytosolic CAS activity is due to cytosolic CS and suggested that the CAS activity of CS is likely to be involved in cyanide metabolism in plant tissues.

A Competitive Enzyme-Linked Immunosorbent Assay to Quantify Acetaldehyde-Protein Adducts That Accumulate in Dry Seeds during Aging.

A competitive enzyme-linked immunosorbent assay (ELISA) was developed to quantify endogenous acetaldehyde-protein adducts (APAs) produced in plant seeds at low acetaldehyde concentrations without exogenous reducing agents. The key point of this technique is the use of a gelatin-acetaldehyde adduct, which is synthesized under 1 mM acetaldehyde and 10 mM NaCNBH3, to pre-coat plate wells to obtain the proper binding parameters for the quantification of APA in seed proteins. Compared with the traditional, direct ELISA method, the competitive one has higher sensitivity and less background. Using competitive ELISA, we determined the accumulation of endogenous APAs in seeds in relation to the loss of seed viability. Lettuce seeds were exposed to 2 mM gaseous acetaldehyde during storage for 30 or 45 d; the relative humidity and temperature of storage were studied independently. Viability decreased only in acetaldehyde-treated seeds, as either the temperature or the relative humidity increased. A loss in viability was accompanied by an increase in the accumulation of APA. The APA content also increased as viability decreased in five species of seeds, which were aged naturally without exposure to acetaldehyde. It is suggested that the modification of functional seed proteins with endogenously evolved acetaldehyde may be an important cause of seed aging.

Interrelations between Carbon Dioxide and Ethylene on the Stimulation of Cocklebur Seed Germination.

Interrelations between CO(2) and C(2)H(4) on promotion of seed germination were examined in more detail at 23 degrees C with presoaked upper seeds of Xanthium pennsylvanicum Wallr. The germination-promoting effect of C(2)H(4) decreased gradually as its application time was delayed during a soaking period, whereas CO(2) was most promotive in application at 5 days of soaking, then its effect declined. CO(2) and C(2)H(4) were additive in earlier soaking periods and synergistic in later periods. Such changes in germination behavior in response to CO(2) and/or C(2)H(4) during a soaking period were closely associated with growth responsiveness of the axial tissues, but not of the cotyledonary ones. Growth responsiveness of axial tissues to CO(2) or C(2)H(4) disappeared finally during a soaking period, but their extinct responsiveness to any one of these gases was almost fully restored in the simultaneous presence of the other. The extinct responsiveness to CO(2) was partially recovered by a preexposure to C(2)H(4). This suggests that in the later period of soaking, unlike the case in a very early period of soaking, the C(2)H(4)-sensitive phase for seed germination precedes the CO(2)-sensitive phase in which CO(2) potentiated axial growth. The restoration of CO(2) responsiveness in axial growth occurred not only after C(2)H(4) treatment but also after exposure to 8 or 33 degrees C or after KCN treatment. Thus, secondarily dormant Xanthium seeds could germinate in response to CO(2) alone, when they were previously exposed for shortterms not only to C(2)H(4) but also 8 degrees C, 33 degrees C, or KCN.

Cyanide-sensitive and Cyanide-resistant Respiration in the Germination of Cocklebur Seeds.

Interrelation between the CN-sensitive cytochrome path and the CN-resistant, benzohydroxamic acid (BHM)-sensitive, or n-propylgallate (nPG)-sensitive alternative path in seed respiration during germination was examined using the nondormant upper and lower seeds of Xanthium pensylvanicum Wallr. The operation of both paths was required not only for normal germination of the lower seed but also for KCN- or NaN(3)-induced germination of both. From the sensitivity to BHM of the germination response, it became obvious that the alternative path exerts its physiological activity as soon as it develops during the early period of water imbibition. Pretreatments with KCN and NaN(3) for promoting germination, strikingly decreased only the engagement of the cytochrome path in the subsequent respiration without affecting that of the alternative path. Nevertheless, no germination occurred without the operation of the cytochrome path. This suggested that excess operation of the cytochrome path is detrimental to germination, being maximal following the BHM-sensitive phase.The alternative and cytochrome paths operated in a proportion of 1 to 5 in a period just before the lower seeds started to germinate. However, there was little difference between the upper seeds of relatively low germination potential and the lower seeds of relatively high potential with respect to both the developmental pattern of the alternative path and the balance of the alternative and cytochrome paths. The higher germination potential of the lower seeds may be related to their high capacities for the alternative path.

D-Amino-acid-stimulated ethylene production in seed tissues.

Ethylene production by axial and cotyledonary tissues excised from Xanthium pennsylvanicum Wallr. seeds was markedly (up to 5-fold) stimulated by the D-isomers of phenylalanine, valine, leucine, threonine, methionine and eithionine while the L-isomers caused no such effect. Responsiveness of these seed tissues to D-methionine appeared soon after the beginning of imbibition, reached a maximum after 6-12 and 12-24 h for the axial and cotyledonary tissues, respectively, and then decreased sharply. D-Phenylalanine and D-methionine also stimulated ethylene production in seed tissues of X. canadense Mill. and in cotyledonary segments from seeds of Helianthus annuus L., Cucurbita moschata Duch. and Vigna radiata (L.) Wilczek. The endogeneous ethylene production and the D-amino-acid-stimulated ethylene production by the seed segments was strongly inhibited by aminoethoxyvinyl glycine, a potent inhibitor of ethylene synthesis from L-methionine.

Possible Involvement of the Alternative Respiration System in the Ethylene-stimulated Germination of Cocklebur Seeds.

Respiration of nondormant upper cocklebur (Xanthium pensylvanicum Wallr.) seeds was enhanced by exogenous C(2)H(4), proportionally to the concentration of C(2)H(4) and the duration of presoaking of the seeds. Benzohydroxamic acid (BHM) and salicylhydroxamic acid (SHM), inhibitors of alternative respiration, inhibited both the germination of nondormant lower cocklebur seeds and the respiration of the upper seeds presoaked for periods of 12 to 30 hours. Both the growth and respiration of axial and cotyledonary tissues were also inhibited by BHM. Moreover, BHM inhibited both the C(2)H(4)-induced germination of the upper seeds and their C(2)H(4)-stimulated respiration; the inhibition occurred only with concomitant addition of C(2)H(4) and BHM. The respiration of seeds with a secondary dormancy induced by presoaking for prolonged periods was markedly stimulated by C(2)H(4) but not suppressed by BHM. It was suggested that the alternative respiration system may be involved in the normal germination process of cocklebur seeds, secondary dormancy may result from its inactivation, and C(2)H(4) may exert its germination-promoting action by stimulating the alternative respiration. The effects of BHM and SHM can suggest but not prove the involvement of the alternative respiration in seed germination.

Thermoperiodism in cocklebur seed germination.

Germination potential in nondormant, upper cocklebur (Xanthium pensylvanicum Wallr.) seeds, which were incapable of germinating under constant temperatures below 25 C in air, was increased by exposure to diurnally alternating temperatures. The cocklebur seeds failed to respond to the temperature fluctuations in the beginning of water imbibition, and their responsiveness appeared only after aerobic presoaking for a limited period or after anaerobic pretreatment for 1 to 3 days.Maximal germination was obtained after exposure to a thermoperiodic regime of 8 hours at 23 C and 16 hours at 8 C. A process occurring during the high temperature phase was aerobic and had to precede the inductive low temperature phase, its effect increasing with temperature. Critical minimum length of the inductive low temperature phase changed with the duration of a preceding anaerobiosis, for instance about 4 hours after 1 day anaerobiosis, but about 2 hours after 2 days. Percentage of subsequent germination was in proportion to the number of thermoperiodic cycles. A process of the inductive low temperature phase was not perturbed by inserting a brief higher temperature period into its phase; indeed, such insertion rather increased germination potential when performed in the earlier parts of the inductive low temperature phase. The effect of the low temperature survived for 13 to 17 hours during the higher temperature period.

Dormancy and Impotency of Cocklebur Seeds: IV. Effects of Gibberellic Acid, Benzyladenine, Thiourea, and Potassium Nitrate on the Growth of Embryonic Axis and Cotyledon Segments.

Germination of nondormant but impotent small cocklebur seeds (Xanthium pennsylvanicum Wallr.) was promoted profoundly with thiourea or benzyladenine, and slightly with gibberellic acid. Gibberellic acid was ineffective in causing the germination of dormant cocklebur seeds, although thiourea and benzyladenine were effective. Experiments with excised seed pieces showed that the promotive effects of thiourea, benzyladenine, and gibberellic acid on cocklebur seed germination were associated with the enhancement of growth of seed parts; thiourea stimulated predominantly the axial growth, whereas benzyladenine stimulated predominantly the cotyledonary growth.Potassium nitrate or indoleacetic acid had little effect on the initial growth of either axes or cotyledons. Except for gibberellic acid, all of the compounds employed enhanced ethylene production, but in general, the ethylene production seemed more likely to be a consequence of growth rather than a cause of it. We concluded that the chemical regulation of seed germination may be a consequence of the alteration of growth capabilities in either the axes or cotyledons, or both.

Dormancy and Impotency of Cocklebur Seeds: VII. Inability of Dormant Cotyledons to Form Chlorophyll.

Dormant seeds of cocklebur (Xanthium pennsylvanicum Wallr.) were characterized by the lack of ability to form chlorophyll. Such an inability of cotyledons of the dormant seeds was improved by the application of various factors and reagents which were capable of breaking the dormancy and of increasing cotyledon enlargement. Of these, ethylene, benzyladenine, and high temperature treatments were particularly effective, and, in turn, oxygen enrichment, gibberellic acid, thiourea, carbon doxide, and potassium nitrate were also promotive to the greening of the dormant cotyledons. The effects of benzyladenine, oxygen enrichment, and high temperature were reduced in the presence of absorbents for endogenously evolve carbon dioxide and ethylene. delta-Aminolevulinic acid could not restore their greening ability.

Gibberellin-induced inhibition and promotion of sprouting in aerial tubers of Begonia evansiana Andr. in relation to photoperiodic treatment and tuber stage.

Gibberellic-acid (GA3) treatment, when applied within a period ranging from the start of short-day (SD) treatment until about 10 SD, GA3 strongly inhibited formation of aerial tubers in response to SD and brought about sprouting of developing aerial tubers. In contrast, when applied after about 10 SD or more, GA3 hastened the completion of the dormant state in the tubers and prolonged their dormancy. The dormancy-promoting effect of GA3 on detached tubers increased with their degree of maturation. Application of growth retardants N-dimethylaminosuccinamic acid (B-9), 2-isopropyl-4-dimethylamino-5-methylphenyl-1-piperidine carboxylate methyl chloride (AMO-1618) and 2-chloroethyltrimethylammonium chloride (CCC) to the cuttings delayed the onset of dormancy in the aerial tuber. When the retardants were applied to detached aerial tubers, however, such a delay of dormancy was not observed, and GA3 application did not inhibit sprouting in aerial tubers detached from CCC-treated cuttings.

Aging progression involving dwarfism and its acceleration by red light in bean hypocotyls.

The effect of red light on the aging progression of the bean (Phaseolus vulgaris L.) hypocotyl segment unit was examined in relation to dwarfism using Kentucky Wonder (tall) and Masterpiece (dwarf) varieties. In both plants, red light promoted the elongation of younger zones and inhibited that of mature zones. The zone exhibiting maximum elongation was shifted to the younger zones by red light irradiation regardless of the plant type, but its extent was greater in the dwarf than in the tall. Thus, red light hastens both the beginning of elongation in the younger portion and its termination in the mature portion of the hypocotyl, particularly of the dwarf plant. These red light responses in each zone of both the tall and dwarf hypocotyl units were reversed by subsequent exposure to far red light regardless of the duration and intensity of red light, thus indicating that the hastened aging progression of the hypocotyl by red light is mediated by phytochrome. However, there is no difference in the rate of decay of Pfr between the tall and dwarf hypocotyls.The increased expression of bean dwarfism seems to result from a concerted action of red light upon the maturation of younger portions of the hypocotyls, which is more rapid in the dwarf than in the tall, and upon the elongation of the relatively matured portion, which is more severely inhibited in the dwarf.

Two C2H 4-producing systems in cocklebur seeds.

C2H4 production of the embryonic axes and cotyledons excised from dormant and non-dormant cocklebur (Xanthium pennsylvanicum Wallr.) seeds was examined in relation to ambient O2 tensions. There were two kinds of C2H4-producing systems, quasi-anaerobic and aerobic, in both organs. Regardless of the organ, the former activity was high in the dormant state and, particularly in axes, declined with after-ripening. On the other hand, the latter activity was almost insignificant in the dormant state, but increased with release from dormancy and the non-dormant axes exclusively produced C2H4 through this system. In the cotyledons, however, the former was still predominant even after they were fully after-ripened. Thus, the C2H4-producing systems were different in the seed organ and in the dormancy state.

Induction of cocklebur seed germination by anaerobiosis: A question about the "inhibitor hypothesis" of seed dormancy.

Germination of non-dormant small cocklebur (Xanthium pennsylvanicum Wallr.) seeds was improved by immersing them in water, suggesting that during their germination endogenous germination inhibitors are leached out. However, the same effect could be obtained by the quasi-anaerobic pre-incubation of the seeds. When seeds were fully imbibed, moreover, water immersion could no longer potentiate them to germinate, and only anaerobiosis increased the germination potential, thus raising a question against the "inhibitor hypothesis" of seed dormancy.

Differential hormone responses in different growing zones of the bean hypocotyl.

Differential growth responses to plant growth substances-a cytokinin (benzyl adenine), gibberellic acid (GA3), indoleacetic acid (IAA), abscisic acid (ABA), coumarin and ethylene-and to environmental factors (carbon dioxide and red light) were found in tissues of different age in a 4-cm segment of the etiolated hypocotyl of Phaseolus vulgaris L. Elongation in all zones of the hypocotyl which were growing or were about to grow was generally enhanced, but each factor or phytohormone acted specifically on a particular zone, in the order cytokinin, red light, GA3, CO2, C2H4 and IAA, going from the top of the hypocotyl. ABA inhibited elongation both in the red-light- and GA3-dependent, immature zones and in the IAA-dependent, relatively mature zones. The effects of C2H4 and CO2 were opposite and those of auxin and coumarin were similar.

Effects of Oxygen and Respiratory Inhibitors on Induction and Release of Dormancy in Aerial Tubers of Begonia evansiana.

Effects of O(2) and some respiratory inhibitors on the induction and release of bud dormancy were examined with the aerial tubers of different ages of Begonia evansiana Andr. Oxygen was needed not only for tuber sprouting but also during the chilling process at 2 to 5 C to break tuber dormancy. If the mature tubers were exposed to blue light during the chilling period, their dormancy was strikingly released even by the chilling given under an O(2) concentration as low as 3%. Blue light pretreatment promoted photo-sprouting of immature tubers only when given under lower O(2) concentrations. On the other hand, red light became effective in inducing dormancy in the immature tubers and in prolonging dormancy in the mature tubers as O(2) tension was increased. This was also the case with the induction of dormancy in the immature tubers by exposing them to a lower temperature (17 C) in the dark. The development of dormancy was suppressed by 2,4-dinitrophenol, p-nitrophenol, and sodium azide.

Dormancy regulation by morphactin in aerial tubers of Begonia evansiana.

The sprouting of aerial tubers of Begonia evansiana was promoted by treatment with morphactin. As with cytokinins, the promotion of sprouting occurred in both the immature and mature tubers. Unlike cytokinins, however, morphactin did not stimulate tuber enlargement. The sprout-inhibiting action of applied gibberellin (GA) was overcome by morphactin. The possible mechanism of the inhibitory action of GA is diseussed in relation to apical dominance.