Absence of fluence rate dependency of phytochrome modulation of stem extension in light-grown Sinapis alba L.

In background white light, supplementary far-red (_max 700 nm) is an order of magnitude less effective than supplementary far-red (_max 739 nm) in the stimulation of stem extension in Sinapis alba. The relationship between phytochrome photoequilibrium and extension rate increase for the two supplementary far-red treatments is, however, very similar. This evidence indicates that phytochrome cycling is not involved in the phytochrome control of stem extension in light-grown Sinapis alba and that the response to supplementary far-red light is not fluence rate (irradiance) dependent.Abbreviations Pfr far-red absorbing form of phytochrome - the phytochrome photoequilibrium (Pfr/Ptotal)     

The influence of light quality on the phytochrome content of light grown Sinapis alba L. and Phaseolus aureus Roxb.

The effect on the phytochrome system of light regimes establishing a range of photoequilibria was studied in two light grown dicotyledonous plants, both of which were treated with the herbicide SAN 9789 to prevent chlorophyll accumulation. In Sinapis alba L. cotyledons the results are comparable with phytochrome behaviour in etiolated mustard seedlings; the level of Pfr becomes independent of wave-length whereas the total phytochrome level is wave-length dependent. Contrasting properties are exhibited in Phaseolus aureus Roxb. leaves in which total phytochrome is unaffected by light quality; consequently the Pfr level is dependent on wavelength. Nevertheless, the amount of phytochrome in mung leaves increased after transfer to darkness suggesting that light still has a profound influence on the phytochrome system, even though light quality during the light period and prior to darkness does not.Abbreviations FR far-red light - WL white light - PAR photosynthetically active radiation - Pfr far-red light absorbing form of phytochrome - Pr red light absorbing form of phytochrome - Ptot total phytochrome level (=Pr+Pfr) - Pfr/Pfr+Pr - SAN 9789 4-chloro-5-(methylamino) 2(,, trifluoro-m tolyl)-3(2H)-pyridazinone     

Rapid photomodulation of stem extension in light-grownSinapis alba L.

Treatment of the whole of aSinapis alba plant with supplementary far-red light (FR), in back-ground white light (WL), induces a rapid increase in stem extension rate. This rapid increase is regulated by the light environment of the stem itself. Supplementary FR to the stem increases extension rate after a lag period of 10–15 min. A lag period of 3–4 h follows FR irradiation of the leaf, before an increase in extension rate is detectable. When the stem is given supplementary FR, the change in extension rate which is induced increases with increasing FR fluence rate, and with decreasing phytochrome photoequilibrium. There is no difference between the effects of supplementary FR _max 719 nm and supplementary FR _max 739 nm for these relationships. The increase in extension rate induced by supplementary FR is reversed by an increase in the fluence rate of red light (R). These data indicate that the response is controlled by phytochrome photoequilibrium.Abbreviations B blue light - FR far-red light - R red light - WL white light - Pfr far-red absorbing form of phytochrome - Pr red absorbing form of phytochrome - P_tot total phytochrome level (=Pr+Pfr); -Pfr/Ptot, measured - ER difference in stem extension rate, before and after treatment     

Amino-acid biosynthesis in the cotyledons of Sinapis alba L. in darkness and far-red light studied by deuterium labelling and mass spectrometry

The incorporation of deuterium from deuterium oxide into the free amino acids of the cotyledons of Sinapis alba L. was studied by gas chromatography-mass spectrometry and was similar, both qualitatively and quantitatively, after incubation of the seedlings in darkness or far-red light. The results support studies which show that phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is synthesised de novo, rather than activated, in response to far-red light.Abbreviations GC-MS Gas chromatography-mass spectrometry - PAL phenylalanine ammonia-lyase (EC 4.3.1.5) - HFB n-propyl heptafluorobutyryl n-propyl     

High irradiance response promotion of a subsequent light induction response in Sinapis alba L.

Relative quantum responsivity curves for inhibition of hypocotyl elongation in Sinapis alba L. seedlings previously grown in white light confirm that a marked end of day inhibition response can be induced by a monochromatic light treatment (30 min) at the end of the light period. In dark grown seedlings, however, no growth inhibition can be induced by a 30 min monochromatic light treatment. A prerequisite for an induction response appears to be a pretreatment with continuous light. Far red light is most effective with blue and red light showing a lesser effectiveness. The light pretreatment also shows a marked fluence rate dependency with respect to its ability to allow an induction response to manifest itself. The pretreatment required shows all the characteristics of a classical HIR response. The appearance of the effect in plants treated with the herbicide SAN 9789 seems to exclude chlorophyll as being the photoreceptor.Abbreviations SAN 9789 4-chloro-5-(methylamino)-2-(, , -trifluoro-m-tolyl)-3(2H)-pyridazinone - RG9 light long wavelength far red light (Schott RG9 colour glass) - FR far red light - WL white light - BL blue light - RL red light - D darkness - P_tot total phytochrome - P_fr far red absorbing form of phytochrome - HSR high irradiance response     

Light-controlled inhibition of hypocotyl growth inSinapis alba L. seedlings

Fluence rate-response curves were determined for the inhibition of hypocotyl growth in 54 h old dark-grownSinapis alba L. seedlings by continuous or hourly 5 min red light irradiation (24 h). In both cases a fluence rate-dependence was observed. More than 90% of the continuous light effect could be substituted for by hourly light pulses if the total fluence of the two different light regimes was the same. Measurements of the far red absorbing form of phytochrome ([P _fr]) and [P _fr]/[P tot] (total phytochrome) showed a strong fluence rate-dependence under continuous and pulsed light which partially paralleled the fluence rate-response curves for the inhibition of the hypocotyl growth.Abbreviations R red - HIR high irradiance response - P _rfr phytochrome in its red, far-red absorbing form - [P _tot]=[P _r]+[P _fr] =k ₁/(k ₁+k ₂): photoequilibrium of phytochrome at wavelength , wherebyk _1,2 rate constants ofP _rP _fr,P _frP _r photoconversion - [P _fr]/[P _tot]     

Phytochrome control of plastid mRNA in mustard (Sinapis alba L.)

The steady-state levels of plastid RNA sequences in dark-grown and light-grown mustard (Sinapis alba L.) seedlings have been compared. Total cellular RNAs were labeled in vitro with ³²P and hybridized to separated restriction fragments of plastid DNA. Cloned DNA fragments which encode the large subunit (LS) of ribulose-1,5-bisphosphate carboxylase [3-phospho-D-glycerate carboxylase (dimerizing), EC 4.1.1.39] and a 35,000 plastid polypeptide were used as probes to assess the levels of these two plastid mRNAs. The 1.22-kilobase-pair mRNA for the 35,000 polypeptide is almost undetectable in dark-grown seedlings, but is a major plastid mRNA in light-grown seedlings. The hybridization analysis of RNA from seedlings which were irradiated with red and far-red light indicates that the level of this mRNA, but not of LS mRNA, is controlled by phytochrome.Abbreviations LS large subunit - RuBP ribulose-1,5-bisphosphate - ptDNA plastid DNA     

Analysis of light-controlled accumulation of carotenoids in mustard (Sinapis alba L.) seedlings

Carotenoid accumulation in the cotyledons of the mustard seedling (Sinapis alba L.) is controlled by light. Besides the stimulatory function of phytochrome in carotenogenesis the experiments reveal the significance of chlorophyll accumulation for the accumulation of larger amounts of acrotenoids. A specific blue light effect was not found. The data suggest that light exerts its control over carotenoid biogenesis through two separate mechanisms: A phytochrome regulation of enzyme levels before a postulated pool of free carotenoids, and a regulation by chlorophyll draining the pool by complex-formation.Abbreviations Chl chlorophyll(s) - PChl protochlorophyll(ide) - HIR high irradiance reaction (of phytochrome) - P_fr far-red absorbing, physiologically active form of phytochrome - P_r red absorbing, physiologically inactive form of phytochrome - P_fof total phytochrome, i.e. [P_r]+[P_fr] - [P_fr]/[P_fof], wavelength dependent photoequilibrium of the phytochrome system - red red light - fr far-red light     

Phytochrome action in light-grown mustard: kinetics,fluence-rate compensation and ecological significance

Internode extension in young, light-grown mustard plants was measured continuously to a high degree of resolution using linear voltage displacement transducers. Plants were grown in background white light (WL) and the first internode was irradiated with supplementary far-red (FR) from fibre-optic light guides, depressing the Pfr/P (ratio of FR-absorbing form of phytochrome to total spectrophotometrically assayable phytochrome) within the internode and causing an acceleration of extension rate. The internode was sensitive to periods of FR as brief as 1 min, with a sharp increase in extension rate occurring after the return to background WL only. The mean latent period of the response to FR was approx. 10 min. Periods of FR longer than approx. 35 min caused an apparently biphasic growth response, with an initial sharp acceleration in extension rate (Phase 1) being followed by a brief deceleration and a further acceleration to a more-or-less steady elevated rate, somewhat less than the first peak (Phase 2). With such longer-term FR, extension rate decelerated upon FR switch-off after a mean lag of approx. 6 min, achieving the prestimulation extension rate within 16 min. The magnitude of the FR-induced increase in extension rate, expressed as a percentage of the rate in WL alone, was an inverse, linear function of the phytochrome photoequilibrium (i.e. Pfr/P, measured in etiolated test material irradiated under the same geometry) over the range 0.17 to 0.63. This relationship was not significantly affected by variations in backround WL fluence rate over the range 50–150 mol·m^-2·s^-1 and was held both for Phase 1 and Phase 2 of the response. The data provide evidence for rapid coupling/uncoupling between phytochrome and its transduction chain in the light-grown plant and for fluence-rate compensation of the regulation of extension rate. The extensive linearity of the relationship between phytochrome photoequilibrium and proportional extension rate increment allows for fine tuning in shade avoidance. The results are discussed with respect to recent evidence on the nature of phytochrome in light-grown plants and in relation to the function of phytochrome in plants growing in the natural environment.Abbreviations FR far-red light - LVDT linear voltage displacement transducer - P total spectrophotometrically assayable phytochrome - PAR photosynthetically active radiation (400–700 nm) - Pfr FR-absorbing form of phytochrome - Pr R-absorbing form of phytochrome - R red light - WL white light     

Photocontrol of hypocotyl elongation in light-grown Cucumis sativus L.

The control by phytochrome of hypocotyl elongation of light-grown Cucumis sativus L. after a white-light period was examined. The farred-absorbing form of phytochrome inhibits hypocotyl elongation. The response to phytochrome photostationary state () is not linear; all values of from 0.004 to 0.13 promote growth maximally, in the range of values of from 0.13 to 0.22 there is a linear growth response, between values of of 0.22 and 0.35 there is again no differential effect, and for values above 0.35 there is a strong (near linear) effect of on elongation. A kinetic examination of events following the white-light period shows that the major recovery from the photoperiod requires 8.5 h of darkness. End-of-day far-red treatment produces a very different response pattern, with a minor growth stimulation within 28 min of treatment followed by a major effect after 80 to 90 min. Three hours after far-red treatment there is a transient decline in growth rate which persists for about 2 h. Over the whole time course there is a great stimulation of growth rate compared with the controls. A similar growth-rate pattern also occurs if the end-of-day is 0.48, although the magnitude of the growth stimulation is less. Two components are affected by end-of-day , namely the time at which growth recovers and the subsequent growth rate. In the long term, the latter accounts for most of the differences in elongation growth. The dark recovery when only the hypocotyl is irradiated requires 4 h, but end-of-day far-red treatment reduces this to about 1.5 h. The persistence of the far-red-absorbing form of phytochrome for many hours in darkness in these light-grown plants is also demonstrated.Abbreviations and symbols D darkness - FR far-red light - Pfr far-red-absorbing form of phytochrome - R red light - WL white light (from fluorescent lamps) - photostationary state of phytochrome - _c calculated      

Persistent effects of changes in phytochrome status on internode growth in light-grown mustard: Occurrence,kinetics and locus of perception

Extension growth of the first internode in fully de-etiolated mustard (Sinapis alba L.) seedlings (11–12.5 d old) is under the control of both the current phytochrome photoequilibrium (Pfr/P, ratio of the far-red-absorbing form of phytochrome to total phytochrome) and that established by short (<12 h) pretreatments. Plants were pretreated with either light pulses providing different calculated Pfr/P followed by dark incubations of different durations (a), or with a 12-h period of white light establishing different Pfr/P (b). After the pretreatments, the plants received either light pulses providing different Pfr/P, followed by dark incubations (c), or continuous white light with or without addtional far-red light (d). Thus, four experimental approaches were followed: (a)(c); (a)(d); (b)(c) and (b)(d). Extension growth during the second period (c or d) was not only affected by the current phytochrome status, but also by that established during the pretreatment period (a or b). The results show the existence of a long-term promotion of stem growth which persists after the end of the low Pfr/P pretreatment. This effect is different from the previously reported rapid effect of far-red light added to background white light as follows: (i) the duration of low Pfr/P required to effect a full response is longer (2.5 h); (ii) the duration of the promotion after returning to high Pfr/P is longer (approx. 24 h) and (iii) the locus of perception is mainly in the leaves, rather than the growing internode.Abbreviations FR far-red light - PAR photosynthetically active radiation - Pfr/P ratio between the FR-absorbing form and total phytochrome - R red light - WL white light     

Regulation of the appearance of glutamine synthetase in mustard (Sinapis alba L.) cotyledons by light,nitrate and ammonium

During transformation of mustard seedlings cotyledons from storage organs to photosynthetically competent leaves, a process which occurs during the first 4 d after sowing, total glutamine-synthetase (GS, EC 6.3.1.2) activity increases from zero to the high level usually observed in green leaves. In the present study we have used ion-exchange chromatography to separate possible isoforms of GS during the development of the cotyledons. The approach failed since we could only detect a single form of GS, presumably plastidic GS, under all circumstances tested. The technique of selective photooxidative destruction of plastids in situ was applied to solve the problem of GS localization. It was inferred from the data that the GS as detected by ion-exchange chromatography is plastidic GS.The regulatory role, if any, of light, nitrate and ammonium in the process of the appearance of GS in the developing cotyledons was investigated. The results show that nitrate and ammonium play only minor roles. Light, operating via phytochrome, is the major regulatory factor.Abbreviations c continuous - D darkness - FPLC fast protein liquid chromatography - GS glutamine synthetase (L-glutamate:ammonia ligase, ADP forming, EC 6.3.1.2) - FR far-red light (3.5 W·m^-2) - NF Norflurazon - R red light (6.8 W·m^-2, _R=0.8)) - RG9-light long-wavelength FR (10 W·m^-2, _RG9<0.01) - () Pfr/Ptot=wavelength-dependent photoequilibrium of the phytochrome system     

A comparative study of the responsivity of Sinapis alba L. seedlings to pulsed and continuous irradiation

Anthocyanin formation in 36h dark grown Sinapis alba L. seedlings and inhibition of hypocotyl elongation in 36h and 54h dark grown and 54h and 7 day light grown seedlings in response to continuous red light could be substituted for by hourly 5 min light pulses where the total fluence over the irradiation period is the same. These pulses are partially (36h) or almost totally (54h and 7 day) reversible by subsequent far-red (RG 9) light pulses. In contrast to 654 nm light, hourly light pulses with 552 nm, 449 nm and 715 nm can at best only partially substitute for continuous irradiation. These data are discussed with respect to the commonly used models for the phytochrome high irradiance response.Abbreviations P_tr tar-red absorbing form of phytochrome - SAN 9789 4-chloro-5-(methyl-amino)-2-(,,-trifluoro-m-tolyl)-3(2H)-pyridazinone=Norflurazon - HIR High irradiance response     

Effect of light on the development of glyoxysomal functions in the cotyledons of mustard (Sinapis alba L.) seedlings

The degradation of storage fat in the cotyledons of mustard seedlings is unaffected by phytochrome and photosynthesis (irradiation with continuous red or far-red light from sowing of the seeds) although light imposes a strong constraint on the translocation of organic matter from the cotyledons into the seedling axis. Likewise, the development and disappearance of glyoxysomal enzyme activities (isocitrate lyase, malate synthase, citrate synthase) takes place independently of light. It is concluded that the mobilization of storage fat (fatcarbohydrate transformation) is independent of photomorphogenesis. The surplus of carbohydrate produced from fat in the light seems to be converted to starch grains in the plastids, which function as a secondary storage pool in the cotyledons.Abbreviations CS citrate synthase - ICL isocitrate lyase - MS malate synthase     

The loci of perception for phytochrome control of internode growth in light-grown mustard: Promotion by low phytochrome photoequilibria in the internode is enhanced by blue light perceived by the leaves

Under continuous white light (WL), extension growth of the first internode in Sinapis alba L. was promoted by low red (R): far-red (FR) ratios reaching the stem and-or the leaves. Conversely, the growth promotion by end-of-day light treatments was only triggered by FR perceived by the leaves and cotyledons, while FR given to the growning internode alone was tatally ineffective. Continuous WL+FR given to the internode was also in-effective if the rest of the shoot remained in darkness. Both the background stem growth, and the growth promotion caused by either an end-of-day FR pulse or continuous WL+FR given to the internode, increased with increasing fluence rates of WL given to the rest of the shoot. The increase by WL of the growth-stimulatory effect of low phytochrome photoequilibria in the internode appears to be mediated by a specific blue-light-absorbing photoreceptor, as blue-deficient light from sodium-discharge lamps, or from filtered fluorescent tubes, promoted background stem growth similarly to WL but did not amplify the response to the R:FR ratio in the internode. Supplementing the blue-deficient light (94 mol·m^-2·s^-1) with low fluence rates of blue (<9 mol·m^-2·s^-1) restored the promotive effect of low R:FR reaching the internode.Abbreviations BL blue light - FR far-red light - PAR photosynthetically active radiation - Pfr/P ratio between the FR-absorbing form and total phytochrome - R red light - SOX low-pressure sodium lamp - WL white light Supported by the Consejo Nacional de Investigaciones Cientificas y Técnicas (República Argentina) and the ORS scheme (UK)     

Factors involved in the coordinate appearance of nitrite reductase and glutamine synthetase in the mustard (Sinapis alba L.) seedling

The extent to which the appearances of nitrite reductase (NIR; EC 1.7.7.1) and glutamine synthetase (GS; EC 6.3.1.2) are coordinated was studied in mustard (Sinapis alba L.) seedlings. It was established by immunotitration that the increased activities of NIR and GS in the presence of light and nitrate can be attributed to the de-novo synthesis of enzyme protein. The bulk of the NIR and GS was found in the developing cotyledons. In the absence of nitrate in the growth medium there was no coordinate appearance of NIR and GS. While light strongly stimulated the appearance of GS, the level of NIR was hardly affected and remained low. On the other hand, in the presence of nitrate in the medium the appearances of NIR and GS were strictly coordinated, the GS level being considerably above that of NIR. It is argued that phytochrome-controlled synthesis of GS in the absence of nitrate is part of the mechanism to reassimilate ammonium liberated during proteolysis of storage protein and metabolism of the resulting amino acids, whereas the strictly coordinated synthesis in the presence of light and nitrate indicates the dominance of nitrate assimilation under these circumstances. The fact that the level of GS was always considerably above that of NIR appears to be a safety measure to prevent ammonium accumulation.Abbreviations FR standardized far-red light (3.5 W·m^–2), to drive the high-irradiance reaction of phytochrome - GS glutamine synthetase, EC 6.3.1.2 - NIR nitrite reductase, EC 1.7.7.1 This work was supported by Heidelberger Akademie der Wissenschaften (Forschungsstelle Nitratassimilation).     

Phytochrome and internode elongation in Chenopodium polyspermum L. the light fluence rate during the day and the end-of-day effect

The elongation of the fourth internode of fully green Chenopodium polyspermum L. is strongly stimulated by far-red light (FR) given at the end of the day. The end-of-day effect is more important when the plants had been cultivated for several days with a main light period of 140 Wm^-2 than with a main light period of 85 Wm^-2. There exists a quantitative relationship between the FR end-of-day effect mediated by phytochrome and the value of the light fluence during the day.Abbreviations D darkness - FR far-red light - HWL white light at 140 Wm^-2 - LWL white light at 85 Wm^-2 - PAR photosynthetically active radiation - R red light - WL white light     

Differential regulation by phytochrome of the appearance of plastidic and cytoplasmatic isoforms of glutathione reductase in mustard (Sinapis alba L.) cotyledons

An increase of glutathione reductase (GR; EC 1.6.4.2) activity during the transformation of mustard (Sinapis alba L.) cotyledons from storage organs to photosynthetically competent leaves was previously found to be controlled by light acting via phytochrome (Drumm, H., Mohr, H., Z. Naturforsch. 28c 559–563, 1973). Two isoforms of GR could be separated by disc electrophoresis. In the present study we have applied ionexchange chromatography to separate isoforms of GR during the development of the cotyledons. Furthermore, the technique of in situ photooxidation of plastids was used to distinguish between plastidic and cytoplasmatic isoforms. The isoform GR₂ is the plastidic enzyme, as shown by its sensitivity to photooxidative treatment, while GR₁ is a cytoplasmatic protein not affected by photooxidative treatment of plastids. Both isoforms are promoted by phytochrome but with different time courses. The appearance of GR₁ is independent of the integrity of the plastids, as one might expect. However, unexpectedly, the phytochrome-mediated re-appearance of GR₂ after a photooxidative treatment is much less affected by photooxidative destruction of the plastids, i.e. by the lack of a particular plastidic factor, than was to be expected from previous experience with typical plastidic proteins. An interpretation of this finding must await measurements at the level of GR₂ mRNA.Abbreviations c continuous - D darkness - FR far-red light (3.5 W·m^-2) - FPLC fast protein liquid chromatography - GR glutathione reductase (EC 1.6.4.2) - NF Norflurazon - R fed light (6.8 W·m^-2) - = Pfr/Ptot wavelength-dependent photoequilibrium of the phytochrome system