Ethylene is not involved in the blue light-induced growth inhibition of red light-grown peas

Although the growth of intact plants is inhibited by irradiation with blue light, the growth rate of isolated stem segments is largely unaffected by blue light. We hypothesized that this loss of responsiveness was a result of ethylene production as part of the wounding response. However, we found no interaction between ethylene- and blue light-induced growth inhibition in dark- or red light-grown seedlings of pea (Pisum sativum L.). Inhibition of growth begins in dark-grown seedlings exposed to blue light within 3 min of the onset of blue light, as was known for red light-grown seedlings. By contrast, ethylene-induced inhibition of growth occurs only after a lag of 20 to 30 min or more (dark-grown seedlings) or 60 min (red light-grown seedlings). Also, the inhibition response of red light-grown seedlings is the same whether ethylene is present from the onset of continuous blue-light treatment or not. Finally the spatial distribution of inhibition following blue light was different from that following ethylene treatment.     

Correlation between Polyribosome Level and the Ability to Induce Nitrate Reductase in Dark-grown Corn Seedlings

Nitrate reductase can be induced in excised shoots of 3-day-old dark-grown Zea mays (var. WF9 × M14) seedlings in the absence of light. In contrast, leaves of 10-day-old dark-grown seedlings require a light treatment in order to induce enzymatic activity. Leaves of 10-day-old dark-grown seedlings contain a very low level of polyribosomes while 3-day-old shoots contain a very high level of polyribosomes. There is a gradual loss of polyribosomes from 3 to 10 days and a gradual loss of in vitro protein synthetic activity of the ribosome preparations. The loss of polyribosomes and decrease in their amino acid-incorporating activity correlate positively with the loss of ability to induce nitrate reducase activity as leaves of dark-grown corn seedlings age. These results corroborate and extend our previous results, in that light is not required for nitrate reductase induction per se in leaves of dark-grown seedlings but is required to reactivate the protein synthetic apparatus of older leaves.     

Cytokinin-induced wall extensibility in excised cotyledons of radish and cucumber

The mechanism of cytokinin-induced cell expansion in cotyledons excised from dark-grown seedlings of radish (Raphanus sativus L.) and cucumber (Cucumus sativus L.) was studied. Cotyledons were incubated in dim light with or without 17 micromolar zeatin for periods up to 3 days. Fresh weights and osmotic potentials were measured daily. Cell wall extensibility properties were measured before and after the growth period. Also, experiments in which radish cotyledons were grown in mannitol solutions of various concentrations were performed. Comparisons of growth rates and increases of tissue osmotic potentials (toward zero) during growth without mannitol indicate that wall extensibility increased during the growth period and that this extensibility was enhanced by zeatin.     

Dim-Red-Light-Induced Increase in Polar Auxin Transport in Cucumber Seedlings : I. Development of Altered Capacity, Velocity, and Response to Inhibitors

We have developed and characterized a system to analyze light effects on auxin transport independent of photosynthetic effects. Polar transport of [³H]indole-3-acetic acid through hypocotyl segments from etiolated cucumber (Cucumis sativus L.) seedlings was increased in seedlings grown in dim-red light (DRL) (0.5 μmol m⁻² s⁻¹) relative to seedlings grown in darkness. Both transport velocity and transport intensity (export rate) were increased by at least a factor of 2. Tissue formed in DRL completely acquired the higher transport capacity within 50 h, but tissue already differentiated in darkness acquired only a partial increase in transport capacity within 50 h of DRL, indicating a developmental window for light induction of commitment to changes in auxin transport. This light-induced change probably manifests itself by alteration of function of the auxin efflux carrier, as revealed using specific transport inhibitors. Relative to dark controls, DRL-grown seedlings were differentially less sensitive to two inhibitors of polar auxin transport, N-(naphth-1-yl) phthalamic acid and 2,3,5-triiodobenzoic acid. On the basis of these data, we propose that the auxin efflux carrier is a key target of light regulation during photomorphogenesis.     

Regulation of 5-Aminolevulinic Acid (ALA) Synthesis in Developing Chloroplasts : II. Regulation of ALA-Synthesizing Capacity by Phytochrome

When dark-grown cucumber (Cucumis sativus L.) seedlings previously exposed to white light for 20 hours were returned to darkness, the ability of isolated chloroplasts to synthesize 5-aminolevulinic acid dropped by approximately 70% within 1 hour. The seedlings were then exposed to light, and the synthetic ability of the isolated chloroplasts was determined. Restoration of the synthetic capacity was promoted by continuous white or red light of moderate intensity. Intermittent red light was also effective. Blue and far-red light did not restore the synthetic capability. Blue light given after a red pulse did not enhance the effect of the red light. Far-red light given immediately after each red pulse prevented the stimulation due to intermittent red light. Restoration of the biosynthetic activity by in vivo light treatments was inhibited by cycloheximide indicating the requirement for translation on 80 S ribosomes for the in vivo light response. These findings suggest that the majority of the plastidic 5-aminolevulinic acid synthesis is under phytochrome regulation.     

Genetic analysis of the role of gibberellin in the red light inhibition of stem elongation in etiolated seedlings

Red light causes a reduction in the extension growth of dark-grown seedlings. The involvement of gibberellin in this process was tested by screening a number of gibberellin synthesis and gibberellin response mutants of Pisum sativum L. for the kinetic response of stem growth inhibition by red light. Gibberellin deficient dwarfs, produced by mutant alleles at the Le, Na, and Ls loci, and gibberellin response mutants produced by mutant alleles at the La and Cry², Lka, and Lkb loci were tested. Extension growth of expanding third internodes of dark-grown seedlings was recorded with high resolution using angular position transducers. Seedlings were treated with red light at a fluence rate of 4 micromoles per square meter per second either continuously or for 75 seconds, and the response was measured over 9 hours. With certain small exceptions, the response to the red light treatments was similar in all the mutants and wild types examined. The lag time for the response was approximately 1 hour and a minimum in growth rate was reached by 3 to 4 hours after the onset of the light treatment. Growth rate depression at this point was about 80%. Seedlings treated with 75 seconds red light recovered growth to a certain extent. Red/far-red treatments indicated that the response was mediated largely by phytochrome. The similar responses to red light among these wild-type and mutant genotypes suggest that the short-term (i.e. 9 hour) response to red light is not mediated by either a reduction in the level of gibberellin or a reduction in the level or affinity of a gibberellin receptor.     

Effects of red light on the growth of intact wheat and barley coleoptiles

The final lengths of intact dark-grown coleoptiles vary with species and cultivar. The growth distribution pattern in the apical 25-mm growing zone and the absolute amount of growth in each zone depend on the age and species of the coleoptile. A comparative study of several cultivars of wheat, Triticum vulgare, and barley, Hordeum vulgare, indicates that the growth distribution pattern in 30- to 38-mm coleoptiles varies with the species and cultivar. In barley, there are two patterns of growth distribution among the several cultivars, whereas in wheat, all cultivars exhibit a common zonal growth pattern. The total growth of coleoptiles, initially 30 to 38 mm in length, during a 24-hour dark incubation period is the same in dark-grown coleoptiles as in those irradiated with 3 minutes of red (660 nm) light prior to the incubation period. The growth distribution pattern in the growing zone of this 30- to 38-mm coleoptile is, however, altered by red light. Growth of the apical 5-mm zone is stimulated by red light and the zonal growth 5 to 10 mm below the apex is only slightly affected, whereas growth in the zones 10 to 15 to 20, and 20 to 25 mm below the apex is inhibited. This growth distribution pattern in irradiated coleoptiles changes as the coleoptile increases in length. The response of a zone following exposure to red light is dependent upon the age of the seedlings irradiated. The over-all effect of red light on growth of the intact coleoptile varies with the length of the coleoptile. In young seedling 20 to 29 mm in length, the cells of the coleoptile can compensate for the effects of red light, with the over-all growth of the dark-grown and irradiated coleoptile about the same. As the seedling grows older, the cells of the coleoptile can no longer make up for the effects of red light, and the over-all effect changes from compensation to pronounced inhibition.     

DIFFERENT SPECIES,SUCROSE, AND LIGHT IN PLANT SECTION ELONGATION TESTS

Sections from both dark- and light-grown seedlings of 11 species were used to test responses to IAA (indoleacetic acid), sucrose, and an inhibitor prepared from cabbage seedlings. Variability among species was great; however, results indicate that many species, light-grown as well as dark-grown, could prove useful in bioassays and probably should be investigated. Although elongation of segments from high-intensity-light-grown cabbage and cucumber hypocotyls and oat coleoptiles had essentially stopped by the time of cutting, their growth and response to IAA as sections were considerable. Neither oat coleoptile nor pea internode sections can be considered representative because of differences in responses to sucrose, of dark-grown sections to light, and to an inhibitor prepared from cabbage. Sucrose generally did not stimulate and even inhibited response of most hypocotyls to IAA. Sucrose was absorbed by sections, increasing final dry weight while not affecting elongation. Sucrose reduced the rate of respiratory decay in cabbage and sunflower, but IAA did not affect respiration. Changes in length and fresh weight of cucumber hypocotyl sections were comparable.     

De novo synthesis of phytochrome in pumpkin hooks

Phytochrome becomes density labeled in the hook of pumpkin (Cucurbita pepo L.) seedlings grown in the dark on D₂O, indicating that the protein moiety of the pigment is synthesized de novo during development. Red light causes a rapid decline of the total phytochrome level in the hook of etiolated seedlings but upon return to the dark, phytochrome again accumulates. These newly appearing molecules are also synthesized de novo. Newly synthesized phytochrome in both dark-grown and red-irradiated seedlings is in the red-absorbing form. Turnover of the red-absorbing form is indicated by the density labeling of phytochrome during a period when the total phytochrome level in the hook of dark-grown seedlings remains constant. However, it was not possible to determine whether this results from intracellular turnover or turnover of the whole cell population during hook growth.     

Influence of Age and Light on the Distribution and Development of Nitrate Reductase in Greening Barley Leaves

The relation between leaf age and the induction of nitrate reductase activity by continuous and intermittent light was studied with barley seedlings (Hordeum vulgare L. cv. Club Mariout). In general, nitrate reductase activity declined as the period of growth in darkness was extended beyond 5 days. Maximum activity was found near the leaf tip while activity was lowest in the morphologically youngest tissue near the base of the lamina. Increased activity was observed after continuous illumination of dark-grown seedlings for 24 hours. The increase in activity in response to light was greatly reduced when the dark pretreatment period was extended beyond 8 days. The amount of nitrate reductase activity present in the different sections of the leaf was closely related to the amount of polyribosomes present. The pattern of chlorophyll accumulation closely parallelled that of increases in nitrate reductase activity. The initial lag in the induction of nitrate reductase activity was removed by a 10-minute light treatment 6 hours before placing dark-grown barley seedlings in light. The enzyme was also induced under flashing light with various dark intervals. These induction curves closely resembled those of chlorophyll accumulation under the same conditions. The development of photosynthetic CO₂ fixation follows the same induction pattern in this system. Our results suggest that photosynthetic products may be required for the induction of significant levels of nitrate reductase activity in leaves of dark-grown seedlings, although other light effects may not be discounted.     

Stimulation of cytokinins and chlorophyll synthesis in cucumber cotyledons by triadimefon

Triadimefon [1-(4-chlorophenoxy)-3,3-dimethyl-1-(1,2,4-triazol-1-yl)-2-butanone] changes the morphology and partitioning of dry matter in cucumber ( Cucumis sativus L. cv. National Pickling) seedlings. The dry weights, potassium and cytokinin levels in the cotyledons and roots of the treated seedlings were higher, whereas the hypocotyl weights were lower than the controls. When etiolated intact seedlings or cotyledons excised from triadimefon-pretreated dark-grown seedlings were exposed to light, chlorophyll synthesis in the pretreated cotyledons was stimulated. Triadimefon does not have cytokinin-like activity in the cucumber cotyledon greening bioassay, but appears to induce the plants to produce more cytokinims, probably by stimulating root growth. Hence it is proposed that the stimulation of chlorophyll production by triadimefon in cucumber cotyledons is mediated by maintaining high levels of potassium and cytokinins in the cotyledons.     

Two distinct blue-light responses regulate epicotyl elongation in pea

Blue light induces a long-term suppression of epicotyl elongation in red-light-grown pea (Pisum sativum L.) seedlings. The fluence-response characteristics are bell-shaped, indicating the possibility of two different blue-light responses: a lower fluence response causing suppression and a higher fluence response alleviating the suppression. To determine if two responses are in effect, we have grown pea seedlings under dark conditions hoping to eliminate one or the other response. Under these growth conditions, only the lower fluence portion of the response (suppression of elongation) is apparent. The kinetics of suppression are similar to those observed for the lower fluence response of red-light-grown seedlings. The response to blue light in the dark-grown seedlings is not due to the excitation of phytochrome because a pulse of far-red light large enough to negate phytochrome-induced suppression has no effect on the blue-light-induced suppression. Furthermore, treatment of the dark-grown seedlings with red light immediately prior to treatment with high fluence blue light does not elicit the higher fluence response, indicating that the role of red light in the blue high fluence response is to allow the plant to achieve a specific developmental state in which it is competent to respond to the higher fluences of blue light.     

Mesocotyl growth of etiolated seedlings ofAvena sativa andZea mays in relation to light and diffusible auxin

Diffusible auxin levels were measured in coleoptiles and mesocotyls of dark-grown seedlings ofavena sativa (cv. Spear) andZea mays (cv. Golden Cross Bantam) using theAvena curvature bioassay. The coleoptile tip was confirmed as the major auxin source in etiolated seedlings. Auxin levels were found to decrease basipetally in sequent sections of theAvena coleoptile but not to decrease in apical sections of increasing length. An inhibitor capable of inducing positive curvatures ofAvena test coleoptiles was discovered in diffusates from the mesocotyls of oat and corn seedlings. The amount of this inhibitor was correlated with the cessation of mesocotyl growth of oat seedlings grown in darkness, and with the inhibition of mesocotyl growth of corn seedlings exposed to red light.     

Pre-germination seed-phytochrome signals control stem extension in dark-grown Arabidopsis seedlings.

The developmental pattern of dark-grown Arabidopsis thaliana is dramatically shifted by exposure of the seedlings to light: inhibition of hypocotyl (stem) growth is one of the typical responses. Here, we show that the hypocotyl growth of dark-grown seedlings is reduced by exposure of the seeds to light. The light signal is perceived by phytochromes A and B during the hours immediately prior to seed germination. The effect is obviously selective, as other processes under phytochrome control were not equally affected by the pre-germination light cue. The hypocotyl response persists for two days after termination of the light signal, which is more than the persistence observed when the seedlings themselves receive the light stimulus. Treatment with far-red light, which converts phytochrome to the inactive form, did not reduce the hypocotyl growth response to pre-germination light, indicating that the persistent signal was not active phytochrome itself. We propose that trans-developmental phase signals could help plants to adjust to their environment.     

Metabolic Adaptations of Plant Respiration to Nutritional Phosphate Deprivation

Shoots of the lazy-2 (lz-2) gravitropic mutant of tomato (Lycopersicon esculentum Mill.) have a normal gravitropic response when grown in the dark, but grow downward in response to gravity when grown in the light. Experiments were undertaken to investigate the nature of the light induction of the downward growth of lz-2 shoots. Red light was effective at causing downward growth of hypocotyls of lz-2 seedlings, whereas treatment with blue light did not alter the dark-grown (wild-type) gravity response. Downward growth of lz-2 seedlings is greatest 16 h after a 1-h red light irradiation, after which the seedlings begin to revert to the dark-grown phenotype. lz-2 seedlings irradiated with a far-red light pulse immediately after a red light pulse exhibited no downward growth. However, continuous red or far-red light both resulted in downward growth of lz-2 seedlings. Thus, the light induction of downward growth of lz-2 appears to involve the photoreceptor phytochrome. Fluence-response experiments indicate that the induction of downward growth of lz-2 by red light is a low-fluence phytochrome response, with a possible high-irradiance response component.     

Photobiology of phytochrome-mediated growth responses in sections of stem tissue from etiolated oats and corn

The far-red reversibility of the phytochrome-controlled stimulation of elongation of coleoptile sections by low fluence red light has been characterized in subapical coleoptile sections from dark-grown Avena sativa L., cv Lodi seedlings. The fluence dependence of the far-red reversal was the same whether or not the very low fluence response is also expressed. The capacity of far-red light to reverse the red light-induced response began to decline if the far-red light was given more than 90 minutes after the red irradiation. Escape was complete if the far red irradiation was given more than 240 minutes after the red irradiation. Sections consisting of both mesocotyl and coleoptile tissue from dark-grown Avena seedlings were found to have physiological regulation of the very low fluence response by indole 3-acetic acid and low external pH similar to that seen for sections consisting entirely of coleoptile tissue. The fluence-dependence of the red light-induced inhibition of mesocotyl elongation was studied in mesocotyl sections from dark grown Zea mays L. hybrid T-929 seedlings. Ten micromolar indole 3-acetic acid stimulates the control elongation of the sections, while at the same time increasing the sensitivity of the tissue for the light-induced inhibition of growth by a factor of 100.     

Rapid Suppression of Growth by Blue Light : BIOPHYSICAL MECHANISM OF ACTION

The mechanism of the rapid inhibition of hypocotyl elongation by blue light was investigated in cucumber (Cucumis sativus L.) and sunflower (Helianthus annuus L.) seedlings by measuring the changes in turgor during the response. A special device, based on the resonance frequency principle, was built which permitted simultaneous and continuous measurements of both tissue rigidity (turgor) and growth rate on a single intact hypocotyl. The large decrease in growth rate following blue irradiation was consistently accompanied by a small increase in resonance frequency. This result indicates that blue light inhibits growth by decreasing the yielding properties of the cell walls, resulting in a slight rise in turgor because of the coupling between growth rate and turgor.     

Inhibition of Zeatin-Induced Growth of Cucumber Cotyledons by Sulfite Ions

The inhibitory effects of sulfite ions on zeatin-induced cellexpansion in cotyledons excised from dark-grown seedlings ofcucumber (Cucumis sativus L.) were examined. With 50 µMzeatin the growth rate in white light was about twice that ofthe control. Addition of Na₂SO₃ in the growth medium inhibitedthe zeatin-induced growth of cotyledons. Zeatin-treatment increasedthe osmotic potential in cell sap of cotyledons, while sulfitedecreased it. These treatments had no significant effect onpotassium concentration. Sulfite inhibited the zeatin-inducedincrease in contents of fructose and glucose, but did not affectsucrose content. The relative contents of non-cellulosic constituentsof cell walls fell with the advance of culture. This decreasewas repressed by sulfite, indicating that inhibition of expansiongrowth in cucumber cotyledons by sulfite ions was the resultof alterations in the cell wall structure due to changes inthe cell wall metabolism. (Received June 12, 1984; Accepted October 24, 1984)