Stem phototropism toward blue and ultraviolet light

Positive phototropism is the process through which plants orient their organs toward a directional light source. While the blue light receptors phototropins (phot) play a major role in phototropism toward blue (B) and ultraviolet (UV) radiation, recent research showed that the UVB light receptor UVR8 also triggers phototropism toward UVB. In addition, new details of the molecular mechanisms underlying the activity of these receptors and interaction with other environmental signals have emerged in the past years. In this review, we summarize the current knowledge about hypocotyledoneous and inflorescence stem growth reorientation toward B and UVB, with a focus on the molecular mechanisms.     

Gibberellic acid and the light inhibition of stem elongation

Summary The ability of gibberellic acid (GA₃) to prevent the light inhibition of stem elongation in peas was examined for several varieties under a wide range of irradiation conditions.A saturating dose of GA₃ largely prevented the inhibitory effect of red light on total stem height in Duke of Albany (tall), Alaska (medium) and Meteor (dwarf) although a small, but statistically significant, effect persisted in all varieties after 3 days of light. The growth of the second internode was, however, markedly inhibited by red light even with a saturating dose of GA₃. With gibberellin there was no difference between the effects of continuous red light and 15 minutes per day on height but the second internode was much shorter in the former treatment. The number of internodes present was the same in both cases and, therefore, the upper internodes in continuous light were as long or longer than in the 15-minute treatment. The number of internodes was only slightly fewer in darkness than in light so that, with GA₃, the effect of red light was transient and only the growth of the lower internodes was inhibited. Without GA₃ overall height was less in both red light treatments than in darkness for all three varieties.In blue light, on the other hand, there was no difference depending on whether height or internode length is considered, and even with a saturating dose of GA₃ the growth rate remained depressed in continuous blue light. There was, however, some interaction between blue light and GA₃.Red/far-red reversal experiments showed that in the varieties Alska and Duke of Albany the far-red stimulation of elongation persisted in the presence of a saturating dose of GA₃ while for the dwarf variety Meteor there was a significant interaction between far-red and GA₃.At least a quantitative difference was found between tall and dwarf peas in their response to light. Tall varieties showed a much greater effect of a prolonged exposure to blue and a smaller effect of a short exposure to red than dwarf varieties. Increasing the duration of exposure to red increasingly inhibited the growth of tall varieties. The medium variety Alaska grew to approximately the same height in continuous red and blue light.     

A study of gibberellin homeostasis and cryptochrome-mediated blue light inhibition of hypocotyl elongation

Cryptochromes mediate blue light-dependent photomorphogenic responses, such as inhibition of hypocotyl elongation. To investigate the underlying mechanism, we analyzed a genetic suppressor, scc7-D (suppressors of cry1cry2), which suppressed the long-hypocotyl phenotype of the cry1cry2 (cryptochrome1/cryptochrome2) mutant in a light-dependent but wavelength-independent manner. scc7-D is a gain-of-expression allele of the GA2ox8 gene encoding a gibberellin (GA)-inactivating enzyme, GA 2-oxidase. Although scc7-D is hypersensitive to light, transgenic seedlings expressing GA2ox at a level higher than scc7-D showed a constitutive photomorphogenic phenotype, confirming a general role of GA2ox and GA in the suppression of hypocotyl elongation. Prompted by this result, we investigated blue light regulation of mRNA expression of the GA metabolic and catabolic genes. We demonstrated that cryptochromes are required for the blue light regulation of GA2ox1, GA20ox1, and GA3ox1 expression in transient induction, continuous illumination, and photoperiodic conditions. The kinetics of cryptochrome induction of GA2ox1 expression and cryptochrome suppression of GA20ox1 or GA3ox1 expression correlate with the cryptochrome-dependent transient reduction of GA(4) in etiolated wild-type seedlings exposed to blue light. Therefore we propose that in deetiolating seedlings, cryptochromes mediate blue light regulation of GA catabolic/metabolic genes, which affect GA levels and hypocotyl elongation. Surprisingly, no significant change in the GA(4) content was detected in the whole shoot samples of the wild-type or cry1cry2 seedlings grown in the dark or continuous blue light, suggesting that cryptochromes may also regulate GA responsiveness and/or trigger cell- or tissue-specific changes of the level of bioactive GAs.     

Desensitization by red and blue light of phototropism in maize coleoptiles

The effects of pretreatments with red and blue light (RL, BL) on the fluence-response curve for the phototropism induced by a BL pulse (first positive curvature) were investigated with darkadapted maize (Zea mays L.) coleoptiles. A pulse of RL, giving a fluence sufficient to saturate phytochrome-mediated responses in this material, shifted the bell-shaped phototropic fluence-response curve to higher fluences and increased its peak height. A pulse of high-fluence BL given immediately prior to this RL treatment temporarily suppressed the phototropic fluence-response curve, and shifted the curve to higher fluences than induced by RL alone. The shift by BL progressed rapidly compared to that by RL. The results indicate (1) that first positive curvature is desensitized by both phytochrome and a BL system, (2) that desensitization by BL occurs with respect to both the maximal response and the quantum efficiency, and (3) that the desensitization responses mediated by phytochrome and the BL system can be induced simultaneously but develop following different kinetics. It is suggested that theses desensitization responses contribute to the induction of second positive curvature, a response induced by prolonged irradiation.Abbreviations BL blue light - RL red light CIW-DPB Publication No. 1001     

Inhibition of stem elongation in cucumis seedlings by blue light requires calcium

The effects of blue light and calcium on elongation of hypocotyl segments of Cucumber (Cucumis sativa L. cv Burpee's Pickler) were studied. Cucumber seedlings grown in dim red light showed a rapid decline in the rate of hypocotyl elongation when irradiated with high intensity (100 micromoles per square meter per second) blue light. In intact, 4-day-old seedlings the inhibition began within 2 minutes after the onset of blue-light irradiation and reached a maximum of approximately 55% within 4 minutes. Hypocotyl segments cut from 4-day-old seedlings also showed an inhibition of elongation in response to blue light when segments were floated on aqueous buffer and exposed to blue light for 3 hours. In the presence of 2 micromolar indole-3-acetic acid, blue light caused a 50% inhibition of elongation. Buffering free calcium in the incubation medium with 0.1 millimolar ethylene glycol bis(-aminoethyl ether)- N,N,N′,N′-tetraacetic acid eliminated the blue-light inhibition of segment elongation. Several experiments confirmed a specific requirement for calcium for the blue-light-induced inhibition of segment elongation. Treating segments with 0.2 micromolar fusicoccin abolished the inhibition of elongation by blue light as did buffering the medium at pH 4. Adding 1 millimolar ascorbate to incubation medium also eliminated the inhibition of segment elongation caused by blue light. Several compounds implicated in cell-wall redox reactions alter the magnitude of the blue-light-induced inhibition. The activity of peroxidase isolated from the cell-wall free space of cucumber hypocotyls was inhibited by ascorbate and low pH. The results are consistent with the hypothesis that blue light inhibits elongation by inducing an increase in cell-wall peroxidase activity and implicate calcium ions in the response to blue light.     

Green light stimulates early stem elongation, antagonizing light-mediated growth inhibition

During the transition from darkness to light, the rate of hypocotyl elongation is determined from the integration of light signals sensed through the phototropin, cryptochrome, and phytochrome signaling pathways. In all light conditions studied, from UV to far-red, early hypocotyl growth is rapidly and robustly suppressed within minutes of illumination in a manner dependent upon light quality and quantity. In this study, it is shown that green light (GL) irradiation leads to a rapid increase in the growth rate of etiolated Arabidopsis seedlings. GL-mediated growth promotion was detected in response to constant irradiation or a short, single pulse of light with a similar time course. The response has a threshold between 10(-1) and 10(0) micromol m(-2), is saturated before 10(2) micromol m(-2) and obeys reciprocity. Genetic analyses indicate that the cryptochrome or phototropin photoreceptors do not participate in the response. The major phytochrome receptors influence the normal amplitude and timing of the GL response, yet the GL response is normal in seedlings grown for hours under constant dim-red light. Therefore, phytochrome activation enhances, but is not required for, the GL response. Seedlings grown under green, red, and blue light together are longer than those grown under red and blue alone. These data indicate that a novel GL-activated light sensor promotes early stem elongation that antagonizes growth inhibition.     

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.     

Hypocotyl growth orientation in blue light is determined by phytochrome A inhibition of gravitropism and phototropin promotion of phototropism

How developing seedlings integrate gravitropic and phototropic stimuli to determine their direction of growth is poorly understood. In this study we tested whether blue light influences hypocotyl gravitropism in Arabidopsis. Phototropin1 (phot1) triggers phototropism under low fluence rates of blue light but, at least in the dark, has no effect on gravitropism. By analyzing the growth orientation of phototropism-deficient seedlings in response to gravitropic and phototropic stimulations we show that blue light not only triggers phototropism but also represses hypocotyl gravitropism. At low fluence rates of blue light phot1 mutants were agravitropic. In contrast, phyAphot1 double mutants grew exclusively according to gravity demonstrating that phytochrome A (phyA) is necessary to inhibit gravitropism. Analyses of phot1cry1cry2 triple mutants indicate that cryptochromes play a minor role in this response. Thus the optimal growth orientation of hypocotyls is determined by the action of phyA-suppressing gravitropism and the phototropin-triggering phototropism. It has long been known that phytochromes promote phototropism but the mechanism involved is still unknown. Our data show that by inhibiting gravitropism phyA acts as a positive regulator of phototropism.     

Primary inhibition of hypocotyl growth and phototropism depend differently on phototropin-mediated increases in cytoplasmic calcium induced by blue light

The phototropin photoreceptors transduce blue-light signals into several physiological and developmental responses in plants. A transient rise in cytoplasmic calcium (Ca2+) that begins within seconds of phototropin 1 (phot1) excitation is believed to be an important element in the transduction pathways leading to one or more of the phot1-dependent responses. The goal of the present work was to determine whether the Ca2+ response was necessary for (a). the inhibition of hypocotyl elongation that develops within minutes of the irradiation, and (b). hypocotyl phototropism (curved growth of the stem in response to asymmetric illumination). After determining that pulses of light delivering photon fluences of between 1 and 1000 micromol m-2 induced growth inhibition mediated by phot1 without significant interference from other photosensory pathways, the effect of blocking the Ca2+ rise was assessed. Treatment of seedlings with a Ca2+ chelator prevented the rise in cytoplasmic Ca2+ and prevented phot1-mediated growth inhibition. However, the same chelator treatment did not impair phot1-mediated phototropism. Thus, it appears that the early, transient rise in cytoplasmic Ca2+ is an important intermediary process in at least one but not all phot1-signaling pathways.     

Root phototropism: from dogma to the mechanism of blue light perception

In roots, the “hidden half” of all land plants, gravity is an important signal that determines the direction of growth in the soil. Hence, positive gravitropism has been studied in detail. However, since the 19th century, the response of roots toward unilateral light has also been analyzed. Based on studies on white mustard (Sinapis alba) seedlings, botanists have concluded that all roots are negatively phototropic. This “Sinapis-dogma” was refuted in a seminal study on root phototropism published a century ago, where it was shown that less then half of the 166 plant species investigated behave like S. alba, whereas 53% displayed no phototropic response at all. Here we summarize the history of research on root phototropism, discuss this phenomenon with reference to unpublished data on garden cress (Lepidium sativum) seedlings, and describe the effects of blue light on the negative bending response in Thale cress (Arabidopsis thaliana). The ecological significance of root phototropism is discussed and the relationships between gravi- and phototropism are outlined, with respect to the starch-statolith-theory of gravity perception. Finally, we present an integrative model of gravi- and blue light perception in the root tip of Arabidopsis seedlings. This hypothesis is based on our current view of the starch-statolith-concept and light sensing via the cytoplasmic red/blue light photoreceptor phytochrome A and the plasma membrane-associated blue light receptor phototropin-1. Open questions and possible research agendas for the future are summarized.     

Spatial separation of light perception and growth response in maize root phototropism

Although the effects of gravity on root growth are well known and interactions between light and gravity have been reported, details of root phototropic responses are less documented. We used high-resolution image analysis to study phototropism in primary roots of Zea mays L. Similar to the location of perception in gravitropism, the perception of light was localized in the root cap. Phototropic curvature away from the light, on the other hand, developed in the central elongation zone, more basal than the site of initiation of gravitropic curvature. The phototropic curvature saturated at approximately 10 micromoles m-2 s-1 blue light with a peak curvature of 29 +/- 4 degrees, in part due to induction of positive gravitropism following displacement of the root tip from vertical during negative phototropism. However, at higher fluence rates, development of phototropic curvature is arrested even if gravitropism is avoided by maintaining the root cap vertically using a rotating feedback system. Thus continuous illumination can cause adaptation in the signalling pathway of the phototropic response in roots.     

Polarity induction versus phototropism in maize: Auxin cannot replace blue light

In a previous study (Nick and Schäfer 1991, Planta 185, 415–424), unilateral blue light had been shown, in maize coleoptiles, to induce phototropism and a stable transverse polarity, which became detectable as stable curvature if counteracting gravitropic stimulation was removed by rotation on a horizontal clinostat. This response was accompanied by a reorientation of cortical microtubules in the outer epidermis (Nick et al. 1990, Planta 181, 162–168). In the present study, this stable transverse polarity is shown to be correlated with stability of microtubule orientation against blue light and changes of auxin content. The role of auxin in this stabilisation was assessed. Although auxin can induce reorientation of microtubules it fails to induce the stabilisation of microtubule orientation induced by blue light. This was even true for gradients of auxin able to induce a bending response similar to that ellicited by phototropic stimulation. Experiments involving partial irradiation demonstrated different perception sites for phototropism and polarity induction. Phototropism starts from the very coleoptile tip and involves transmission of a signal (auxin) towards the subapical elongation zone. In contrast, polarity induction requires local action of blue light in the elongation zone itself. This blue-light response is independent of auxin.This work was supported by the Deutsche Forschungsgemeinschaft and two grants of the Studienstiftung des Deutschen Volkes and the Human Frontier Science Program Organization to P.N.     

Absence of red-light enhancement of phototropism in pea seedlings at limiting irradiances of blue light

The effect of different light qualities (blue, green, white, red and far-red) on ethylene production in leaf discs and flower petal discs of Begonia × hiemalis cv. Schwabenland Red was studied. All the light qualities, except far-red, reduced the ACC-conversion to ethylene in leaf discs by about 70% at a photosynthetic photon flux density (PPFD) of 20 mol m^–2s^–1.Blue and green light were less inhibitory than white and red light at lower PPFD. In all treatments far-red light at 0.5 mol m^–2s^–1 of photon flux density (PFD) stimulated the ACC-conversion to ethylene in leaf discs by about 60–90% compared to the dark-incubated control. White and red light strongly inhibited the -naphthalene-acetic acid (NAA) stimulated ethylene synthesis in leaf discs. The results may suggest that the ethylene production is controlled by phytochrome in the leaves but not in the petals. Lack of coaction of any light quality with silver ions on ethylene production in leaf and petal discs was also observed.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - EFE ethylene forming enzyme - NAA -naphthalene-acetic acid - PFD photon flux density - PPFD photosynthetic photon flux density - RH relative air humidity - SAM S-adenosylmethionine - STS silver thiosulphate     

Specific inhibition of phototropism in corn seedlings

Geotropism was used as a control for the specificity of potential inhibitors of phototropism by the coleoptiles of corn (Zea mays) seedlings. The compounds tested fall into three categories showing: (a) no inhibition of either phototropism or geotropism (KCl); (b) nonspecific inhibition of both phototropism and geotropism (KCN); and (c) specific inhibition of phototropism (KI, NaN₃, and phenylacetic acid). Simultaneous irradiation of coleoptiles with phototropically inert light in addition to the phototropically active blue light also results in an inhibition of phototropism. Since azide, iodide, and phenylacetic acid are known to interact with flavins while a simultaneous irradiation with a phototropically inert light may depopulate the first triplet state of flavins, these data support the hypothesis that the photoreceptor pigment for phototropism in corn is a flavin.     

Regulation of pea epicotyl elongation by blue light : fluence-response relationships and growth distribution

Irradiation with blue light causes a rapid decrease in stem elongation in Pisum sativum. Growing plants under continuous red light allowed us to study the fluence dependence and spatial distribution of blue-induced growth effects without interference from large changes in the ratio of the far-red absorbing form of phytochrome to total phytochrome. The magnitude of the inhibition generated by a 30-second pulse of blue light was linearly related to the log of the fluence applied over two orders of magnitude. Reciprocity held for irradiations with a pulse length shorter than the lag time for the response. The spatial distribution of inhibition was studied by marking the growing zone and photographing the stem at 10-minute intervals before, during, and after a 1-hour exposure to blue light. The region just below the hook does not undergo any perceptible change in growth rate while growth is nearly 100% inhibited in the base of the third internode.     

Involvement of wall microtubules in gibberllin promotion and kinetin inhibition of stem elongation

The elongation of light-grown azuki bean (Azukia angularis =Vigna angularis) epicotyl segments was promoted by indoleaceticacid (IAA) and this IAA-induced elongation was inhibited byboth kinetin and benzimidazole (BIA). Increased stem thickeningwas observed with kinetin- or BIA-treated segments, but thiswas not accompanied by incresed cell number in the transversedirection, suggesting that both kinetin and BIA promoted lateralcell expansion. Colchicine at a concentration with no effecton IAA-induced elongation reversed both the kinetic- and BIA-inducedinhibition. Electron-microscopic examination revealed that wall microtubulesin cells treated with kinetin together with IAA ran parallelto the cell axis, while wall microtubules in cells treated withonly IAA were randomly oriented. In the cell treated with gibberellintogether with IAA, wall microtubules ran tranverse to the cellaxis. (Received July 13, 1973; )     

Genetic loci associated with stem elongation and winter dormancy release in wheat

In winter wheat (Triticum aestivum L.), the stem begins to elongate after the vernalization requirement is satisfied during winter and when favorable temperature and photoperiod conditions are attained in spring. In this study, we precisely measured elongation of the first extended internode on 96 recombinant inbred lines of a population that was generated from a cross between two winter wheat cultivars, Jagger (early stem elongation) and 2174 (late stem elongation). We mapped a major locus for stem elongation to the region where VRN-A1 resides in chromosome 5A. Visible assessment of winter dormancy release was concomitantly associated with this locus. VRN1 was previously cloned based on variation in vernalization requirement between spring wheat carrying a dominant Vrn-1 allele and winter wheat carrying a recessive vrn-1 allele. Both of two winter wheat cultivars in this study carry a recessive vrn-A1 allele; therefore, our results suggest that either VRN-A1 might invoke a new regulatory mechanism or a new gene residing close to VRN-A1 plays a regulatory role in winter wheat development. Phenotypic expression of the vrn-A1a allele of Jagger was more sensitive to the year of measurement of stem elongation than that of the vrn-A1b allele of 2174. In addition to QSte.osu.5A, several loci were also found to have minor effects on initial stem elongation of winter wheat. Seventeen of nineteen locally adapted cultivars in the southern Great Plaints contained the vrn-A1b allele. Hence, breeders in this area have inadvertently selected this allele, contributing to later stem elongation and more conducive developmental patterns for grain production.     

The inhibition of mitosis in tissue culture cells by blue light

Blue light (wavelength 350-480 nm) irradiation of the early mitotic (prophase and prometaphase) tissue culture cells at the dose of 50-3000 J/cm2 delay mitosis or completely block it at the metaphase. Cell sensitivity to the near UV light (wavelength 360 nm) was few times more as compared with the sensitivity to the visible light (wavelength 400-480 nm). Mitotic cells irradiated with the green light (wavelength more than 500 nm; dose up to 7500 J/cm2) completed division normally. The effect of the blue light did not depend on the presence of phenol red in tissue culture medium. Rhodamin 123 staining did not show any changes in the mitochondrial system in the irradiated mitotic cells. Blue light irradiation with the dose enough for the induction of mitotic delay appears to be insufficient to affect the proliferation of interphase cells.     

The blue light receptor Phototropin 1 suppresses lateral root growth by controlling cell elongation

We investigated the relationship between the blue light receptor phototropin 1 (phot1) and lateral root growth in Arabidopsis thaliana seedlings. Fluorescence and confocal microscopy images, as well as PHOT1 mRNA expression studies provide evidence that it is highly expressed in the elongation zone of lateral roots where auxin is accumulating. However, treatment with the auxin transport inhibitor N‐1‐naphthylphthalamic acid significantly reduced PHOT1 expression in this zone. In addition, PHOT1 expression was higher in darkness than in light. The total number of lateral roots was higher in the phot1 mutant than in wild‐type Arabidopsis. Cells in the elongation zone of lateral roots of the phot1 mutant were longer than those of wild‐type lateral roots. These findings suggest that PHOT1 plays a role(s) in elongation of lateral roots through the control of an auxin‐related signalling pathway.     

Evidence from studies with acifluorfen for participation of a flavin-cytochrome complex in blue light photoreception for phototropism of oat coleoptiles

The diphenyl ether acifluorfen enhances the blue light-induced absorbance change in Triton X100-solubilized crude membrane preparations from etiolated oat (Avena sativa L. cv. Lodi) coleoptiles. Enhancement of the spectral change is correlated with a change in rate of dark reoxidation of a b-type cytochrome. Similar, although smaller, enhancement was obtained with oxyfluorfen, nitrofen, and bifenox. Light-minus-dark difference spectra in the presence and absence of acifluorfen, and the dithionite-reduced-minus oxidized difference spectrum indicate that acifluorfen is acting specifically at a blue light-sensitive cytochrome-flavin complex. Sodium azide, a flavin inhibitor, decreases the light-induced absorbance change significantly, but does not affect the dark reoxidation of the cytochrome. Hence, it is acting on the light reaction, suggesting that the photoreceptor itself is a flavin. Acifluorfen sensitizes phototropism in dark-grown oat seedlings such that the first positive response occurs with blue light fluences as little as one-third of those required to elicit the same response in seedlings grown in the absence of the herbicide. Both this increase in sensitivity to light and the enhancement of the light-induced cytochrome reduction vary with the applied acifluorfen concentration in a similar manner. The herbicide is without effect either on elongation or on the geotropic response of dark-grown oat seedlings, indicating that acifluorfen is acting specifically close to, or at the photoreceptor end of, the stimulus-response chain. It seems likely that the flavin-cytochrome complex serves to transduce the light signal into curvature in phototropism in oats, with the flavin moiety itself serving as the photoreceptor.