The in vivo properties of Amaranthus phytochrome

Summary Phytochrome has been measured in etiolated seedling of Amaranthus caudatus. The phytochrome content increases from the time of germination until 72 hr from sowing, after which it remains constant at 27.5x10^-3 (OD) units per 200 seedlings. After a saturating dose of red light P _fr decays in the dark to a form not detectable photometrically. There is no evidence for the process of dark reversion of P _fr to P _fr found in other dicotyledons. Even in the presence of azide, a selective inhibitor of decay, the process of dark reversion is not observed. The decay of P _fr has been investigated at different temperatures and follows first order decay kinetics throughout. Over the temperature range 15–30° the Q ₁₀ of decay remained constant at 4.3.The photostationary states of phytochrome (P _fr /P _total )maintained by mixed red/far-red light have been measured in both seedlings and partially purified protein extracts, with good agreement. The rate of phytochrome decay can be manipulated by changing the P _fr /P _total ratio. The lag period before a decay curve becomes exponential is characteristic of a particular P _fr /P _total ratio and represents the time for attainment of the photostationary state. The effect of energy on decay has been investigated under red and blue light. The rate of phytochrome decay is dependent on the P _fr /P _total ratio and only becomes energy dependent when the light intensity is so low that the photostationary state is never attained.The process of apparent phytochrome synthesis has been found in Amaranthus. After reducing the phytochrome to a low level by red light treatment a rate of apparent synthesis of 1.35×10^-4 (OD) units per hr per 200 seedlings was observed, levelling off at 29% of the original phytochrome level.Under white tungsten lights of high intensity there is a deviation from the expected first order decay kinetics. The nature of this low rate of decay cannot be explained at the present time.     

Kinetics of the dichroic reorientation of phytochrome during photoconversion inMougeotia

In the green algaMougeotia, the dichroic orientation of the red-absorbing form of phytochrome (P_r) is parallel of the cell surface, whereas the far-red-absorbing form (P_fr) is oriented normal to it. The time course of the change from parallel to normal was investigated by double-flash irradiation with polarized red and far-red light. The results obtained by two different methods indicate that most of the phytochrome intermediates existing in the first 5 ms after the inducing red flash are still oriented parallel to the cell surface, similar to P_r. At increasing intervals between the red and the far-red flashes, more and more phytochrome molecules turn their transition moments to the P_fr orientation. This reaction is finished after approximately 30 ms. We conclude that the change in dichroic orientation of the phytochrome molecules inMougeotia occurs during the last relaxation steps of the intermediates on the way from P_r to P_fr. It cannot be decided yet, whether the first surface-normal phytochrome species is an intermediate or P_fr itself.Abbreviations P_r red-absorbing form of phytochrome - P_fr far-red-absorbing form of phytochrome A preliminary report of this work was presented at the European Symposium on Photomorphogenesis, University of Reading, UK (Kraml et al. 1982)     

A Kinetic Theory of First Order Cyclical Processes—Phytochrome Controlled Red Light Induced Cereal Leaf Unfolding Compared with Theory

The phytochrome controlled unfolding of cereal leaves was studied as a function of irradiation time and light intensity (narrowband red light) over a wide energy range (5 decades). With different intensities, a family of similarly shaped response curves appear with distinct time-dependent maxima and minima. A theoretical kinetic model based upon a cyclical phytochrome photoconversion scheme has been calculated by us. The theoretical calculations and the experimental findings are in excellent agreement. The same model explains the early photoresponses (first maxima) as an effect of one active phytochrome form, P₂, and the delayed photoresponses as an effect of a second active form P_n. The active transitory form, P₂ (although it may not be the primary product), is formed upon light absorption from P₁. The P₂ decays by a first order dark reaction through several inactive intermediates to P_n (active). The effect of the intermediates is mainly to delay the production of the second active product. It is possible to identify the two active products, P₂ and P_n, as P_fr and P*_fr, respectively. The presented cyclical phytochrome reaction scheme is a special case of a general first order kinetic cycle which includes all possible feed back loops. The latter scheme also has been calculated and programmed since it has a more general application.     

Aspects of phytochrome decay in etiolated seedlings under continuous Illumination

Summary The rate of total phytochrome decay in the dicotyledons Amaranthus caudatus, Mirabilis jalapa and Pisum sativum under continuous illumination with red, incandescent, and blue light depends on the P_FR/P_total maintained by each source. Amaranthus is an exception to this in that there is a deviation from firstorder decay kinetics under continuous illumination with incancdescent light. This deviation is probably not related to the chlorophyll present in the Amaranthus sample since chlorophyll-rich Pisum buds have the same phytochrome decay rate as epicotyl tissue under continuous incandescent light. Reports of a prolonged lag phase before the onset of first-order decay kinetics of phytochrome in Pisum have not been confirmed and the small lag phase observed in the present work can be accounted for by the time required to attain the P_FR/P_total ratio characteristic of blue light in a carotenoid rich tissue. In the monocotyledon, Avena sativa, and perhaps monocotyledons in general, decay rate is maximal at a low P_FR concentration and the decay curve is the same under continuous red, incandescent and blue light. This dicotyledon/monocotyledon difference with respect to saturation of phytochrome decay does not correlate with the other dicotyledon/monocotyledon difference, the presence or absence of dark reverions of P_FR to P_R, since the dicotyledons Amaranthus and Mirabilis that lack reversion still show no saturation of decay. Possible growth control by the P_FR/P_total ratio is discussed in relation to environmental changes in light quality.Research carried out at Brookhaven National Laboratory under the auspices of the U. S. Atomic Energy Commission.     

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-controlled extension growth of Avena sativa L. seedlings

The effects of continuous red and far-red light and of brief light pulses on the growth kinetics of the mesocotyl, coleoptile, and primary leaf of intact oat (Avena sativa L.) seedlings were investigated. Mesocotyl lengthening is strongly inhibited, even by very small amounts of P_fr, the far-red light absorbing form of phytochrome (e.g., by [P_fr]0.1% of total phytochrome, established by a 756-nm light pulse). Coleoptile growth is at first promoted by P_fr, but apparently inhibited later. This inhibition is correlated in time with the rupturing of the coleoptile tip by the primary leaf, the growth of which is also promoted by phytochrome. The growth responses of all three seedling organs are fully reversible by far-red light. The apparent lack of photoreversibility observed by some previous investigators of the mesocotyl inhibition can be explained by an extremely high sensitivity to P_fr. Experiments with different seedling parts failed to demonstrate any further obvious interorgan relationship in the light-mediated growth responses of the mesocotyl and coleoptile. The organspecific growth kinetics, don't appear to be influenced by P_fr destruction. Following an irradiation, the growth responses are quantitatively determined by the level of P_fr established at the onset of darkness rather than by the actual P_fr level present during the growth period.Abbreviation P_fr far-red light absorbing form of phytochrome     

Involvement of phytochrome and a blue light photoreceptor in UV-B induced flavonoid synthesis in parsley (Petroselinum hortense Hoffm.) cell suspension cultures

Flavonoid synthesis in cell suspension cultures of parsley (Petroselinum hortense Hoffm.) occurs only after irradiation with ultraviolet light (UV), mainly from the UV-B (280–320 nm) spectral range. However, it is also controlled by phytochrome. A P_fr/P_tot ratio of approximately 20% is sufficient for a maximum phytochrome response as induced by pulse irradiation. Continuous red and far red light, as well as blue light, given after UV, are more effective than pulse irradiations. The response to blue light is considerably greater than that to red and far red light. Continuous red and blue light treatments can be substituted for by multiple pulses and can thus probably be ascribed to a multible induction effect. Continuous irradiations with red, far red and blue light also increase the UV-induced flavonoid synthesis if given before UV. The data indicate that besides phytochrome a separate blue light photoreceptor is involved in the regulation of the UV-induced flavonoid synthesis. This blue light receptor seems to require the presence of P_fr in order to be fully effective.Abbreviations HIR high irradiance response - P_fr far red absorhing form of phytochrome - P_tet total phytochrome - UV ultraviolet light     

Phytochrome behaves as a dimer in vivo

Abstract It is well established that phytochrome exists as a dimer in vitro. A comparison of the relative photoequilibrium concentrations of P_rP_r, P_rP_fr and P_frP_fr, with the relative sizes of the P_fr-pools which undergo dark reversion in the intact plant, leads to the hypothesis that phytochrome also exists as a dimer in vivo, This hypothesis is in accordance with kinetic properties of the phytochrome system under continuous irradiation. Additional support for this view is provided by the observation that P_fr-destruction after a red light flash, which should favour the formation of P_rP_fr dimers, is paralleled by a decay of P_r, even if the presence of P_r cycled through P_fr can be excluded. Preliminary observations could indicate an interaction of the subunits of a phytochrome dimer during the process of phototransformation.     

Control by light of hypocotyl growth in de-etiolated mustard seedlings

After sowing, mustard (Sinapis alba L.) seedlings were grown for 48 h in white light (25°C). These fully de-etiolated, green seedlings were used as experimental material between 48 and 72 (84) h after sowing. The question researched was to what extent control by light of hypocotyl elongation is due to phytochrome in these seedlings. It was found that the light effect on hypocotyl growth is very probably exerted through phytochrome only. In particular, we found no indication for the involvement of a specific blue light photoreceptor pigment.Abbreviations HIR high irradiance reaction - P_fr far-red absorbing, physiologically active form of phytochrome - P_r red absorbing, physiologically inactive form of phytochrome - Pot total phytochrome, i.e. [P_r]+[P_fr] - [P_fr]/[P_tot] - red red light - fr far-red light - wl white light - bl blue light - di dichromatic irradiation - l hypocotyl length     

Regulation of enzyme levels by phytochrome in mustard cotyledons: Multiple mechanisms?

Phytochrome controls the appearance of many enzymes in the mustard (Sinapis alba L.) cotyledons. The problem has been whether the effect of phytochrome on the appearance of enzymes in this organ is due to a common initial action of P_fr, e.g. due to the liberation of a second messenger. We have compared the modulation by light (phytochrome) of the appearance of phenylalanine ammonia lyase (PAL)⁺ and ribulosebisphosphate carboxylase (Carboxylase)⁺. PAL becomes detectable in the mustard cotyledons at 27 h after sowing while Carboxylase starts to appear only at 42 h after sowing (starting points, 25° C). The starting points cannot be shifted by light. As a major result, in the case of PAL the inductive effect of continuous red light (given from the time of sowing) remains fully reversible by 756 nm-light up to the starting point (27 h after sowing) while with Carboxylase full reversibility in continuous red light is lost at approximately 15 h after sowing. While the induction of Carboxylase is already saturated at a very low level of P_fr (e.g. continuous 756 nm-light saturates the response) and does not depend on irradiance (e.g. continuous 675 mW m^-2 red light and 67.5 mW m^-2 red light lead to the same time course), PAL induction is a graded response over a wide range of P_fr doses and depends strongly on the fluence rate (high irradiance response, HIR). It is concluded that PAL induction and Carboxylase induction are not only separated in time but differ in every regard except that both responses are mediated by phytochrome.The present data support the previous conclusion that the specification of the temporal and spatial pattern of development is independent of phytochrome even though the realization of the pattern of development can only occur in the presence of phytochrome (P_fr). It seems that there is no feedback from pattern realization to pattern specification.Abbreviations P_fr the far-red absorbing, physiologically active form of phytochrome - P_r the red absorbing physiologically inactive form of phytochrome - P_total [P_r]+[P_fr] - PAL phenylalanine ammonia-lyase (EC 4.3.1.5) - Carboxylase ribulosebisphosphate carboxylase (EC 4.1.1.39)     

Nature of light requirement for the flowering of Chenopodium rubrum L. (Ecotype 60° 47′ N)

Seedlings of C. rubrum were irradiated with different light qualities and intensities following a single inductive dark period. Our results show that relatively low intensity white light (35–100 ft. c.) does not support flower development while high intensity white light (650–800 ft. c.) permits 100% flowering. We have shown that the low intensity light inhibiton of flower development is not due to suboptimal photosynthesis. Relatively low intensities of light rich in far-red or blue wavebands sustains optimum flower development, whereas red light is totally ineffective in this respect. Considering that the intensity dependent High Energy Reaction (HER) has its action maxima in the blue and far-red we propose that HER may be positively involved in the flower development of C. rubrum. Our study further suggests that there may be some flower inhibitory component at play in relatively low intensity white light conditions and HER may be required to counteract this flower inhibitory effect.Abbreviations SD short day plant - HER High Energy Reaction - P_FR far-red absorbing form of phytochrome - P_R red absorbing form of phytochrome - L.I.I. low intensity incandescent white light - H.I.I. high intensity incandescent white light - L.I.F. low intensity fluorescent white light - H.I.F. high intensity fluorescent white light - GA₃ gibbrellic acid This paper constitutes a part of a Ph.D. thesis submitted to the University of Western Ontario, London, Ontario.     

Coaction of three factors controlling chlorophyll and anthocyanin synthesis

In a three-factor analysis the rate of chlorophyll a (Chl) accumulation in excised mustard cotyledons was studied as a function of kinetin, light (operating through phytochrome, P _fr) and an excision factor. It was found that the three factors operate additively provided that the P _fr level is high enough. When the P _fr level is below approximately 1 per cent (<0.01) the effectiveness of the excision factor decreases while the effect of kinetin remains additive. The observed additivity is explained by a model where the three factors operate independently through a common intermediate (presumably 5-aminolevulinate) in the biosynthetic chain leading to Chl. With regard to the coaction of the excision factor and phytochrome it is concluded that the production of the excision factor requires the operation of phytochrome (even though saturated at a low P _fr level) while the action of the excision factor is independent of phytochrome. This conclusion was confirmed by experiments in which the rate of light-mediated anthocyanin synthesis was measured in excised mustard cotyledons. The effect of excision in the case of anthocyanin formation differs kinetically from the effect of excision on Chl formation.Abbreviations Chl chlorophyll(ide) a - P _fr far-red absorbing form of phytochrome - P _fr/P _tot ratio at photoequilibrium - RL red light - FR far-red light - GL green light - RG9 light long wavelength far-red light - WL white light     

Light requirement,phytochrome and photoperiodic induction of flowering of Pharbits nil Chois

The low chlorophyll content of cotyledons of Pharbitis nil grown for 24 h in far-red light (FR) or at 18° C in white light from fluorescent lamps (WL) allows spectrophotometric measurement of phytochrome in these tissues. The (A) measurements utilize measuring beams at 730/802 nm and an actinic irradiation in excess of 90 s. The constancy of the relationship between phytochrome content and sample thickness confirms that, under these conditions of measurement, a true maximum phytochrome signal was obtained. These techniques have been used to follow changes in the form and amount of phytochrome during an inductive dark period for flowering. Following exposure to 24h WL at 18° C with a terminal 10 min red (R), P_fr was lost rapidly in darkness and approached zero in less than 1 h; during this period there was no change in the total phytochrome signal. Following exposure to 24 h FR with a terminal 10 min R, P_fr approached zero in 3 h, and the total phytochrome signal decreased by about half. The relevance of these changes to photoperiodic time measurement is discussed.Abbreviations BCJ irradiation from photographic ruby-red lamps - FR far-red light - P_fr far-red-absorbing form of phytochrome - P_r red-absorbing form of phytochrome - P total phytochrome content - R red light - WL white light from fluorescent lamps     

Auxin inhibition of acid-and fusicoccin-induced elongation in lentil roots

Seedlings of the short-day plant, Chenopodium rubrum L. (Ecotype 60° 47 N) were irradiated with different intensities and qualities of light for 24 h preceding a single inductive dark period (12 h). Our data shows that a relatively low intensity incandescent light (35–100 ft. c.) is not effective as the photoperiod for flowering. The above effect is not due to a requirement for a relatively high level of photosynthesis. Our results suggest a definite promotory role of a blue High Energy Reaction (HER). We could not demonstrate the involvement of a far-red HER. We suggest that ineffectiveness of far-red may have been due to establishment of rather low Phytochrome, P _FR , levels, suboptimal for flowering. A certain critical level of P _FR (30–40%, that presumably established by blue light) seems to be necessary for photoreactions involved in flowering of C. rubrum. There are indications in our experiments of the operation of a red radiation mediated flower inhibitory photoreaction.Abbreviations SD short day plant - HER High Energy Reaction - P _FR far-red absorbing form of phytochrome - P _R red absorbing form of phytochrome - L.I.I. low intensity incandescent white light - H.I.I. high intensity incandescent white light - L.I.F. low intensity fluorescent white light - H.I.F. high intensity fluorescent white light - DCMU 3(2, 3, dichlorophenyl) 1, 1 dimethyl urea This paper constitutes a part of a Ph.D. thesis submitted to the University of Western Ontario, London, Ontario     

Variation in dynamics of phytochrome A in Arabidopsis ecotypes and mutants

Phytochromes are photoreceptors in plants which can exist in two different conformations: the red light‐absorbing form (P_r) and the far‐red light‐absorbing form (P_fr), depending on the light quality. The P_fr form is the physiologically active conformation. To attenuate the P_fr signal for phytochrome A (phyA), at least two different mechanisms exist: destruction of the molecule and dark reversion. Destruction is an active process leading to the degradation of P_fr. Dark reversion is the light‐independent conversion of physiologically active P_fr into inactive P_r. Here, we show that dark reversion is not only an intrinsic property of the phytochrome molecule but is modulated by cellular components. Furthermore, we demonstrate that dark reversion of phyA may be observed in Arabidopsis ecotype RLD but not in other Arabidopsis ecotypes. For the first time, we have identified mutants with altered dark reversion and destruction in a set of previously isolated loss of function PHYA alleles (Xu et al. Plant Cell 1995, 7, 1433–1443). Therefore, the dynamics of the phytochrome molecule itself need to be considered during the characterization of signal transduction mutants.     

Kinetics of phytochrome decay in Amaranthus seedlings

Summary In Amaranthus seedlings the disappearance of the unstable P _fr form of phytochrome does not involve dark reversion to P _r . The rate constant for the decay of total phytochrome under continuous illumination is directly related to the proportion in the P _fr form. This relationship allows calculations to be made of the proportion of P _fr under continuous far-red illumination where the amount is too low to be measured directly.     

The role of phytochrome in photoperiodic time measurement and its relation to rhythmic timekeeping in the control of flowering in Chenopodium rubrum

Summary To follow changes in the status of phytochrome in green tissue and to relate these changes to the photoperiodic control of flowering, we have used a null response technique involving 1.5-min irradiations with mixtures of different ratios of R and FR radiation.Following a main photoperiod of light from fluorescent lamps that was terminated with 5 min of R light, the proportion of P_fr in Chenopodium rubrum cotyledons was high and did not change until the 3rd hour in darkness; at this time, P_fr disappeared rapidly. When the dark period began with a 5-min irradiation with BCJ or FR light to set the proportion of P_fr low P_fr gradually reappeared during the first 3 h of darkness and then disappeared again.The timing of disappearance of P_fr is consistent with the involvement of phytochrome in photoperiodic time measurement. Reappearance of P_fr after an initial FR irradiation explains why FR irradiations sometimes fail to influence photoperiodic time measurement or only slightly hasten time measurement. A R light interruption to convert P_r to P_fr delayed, the timer by 3 h but only for interruptions after and not before the time of P_fr disappearance. Such 5-min R-light interruptions did not influence the operation of the rhythmic timekeeping mechanism. Continuous or intermittent-5 min every 1.5 h-irradiations of up to 6 h in duration were required to rephase the rhythm controlling flowering. A skeleton photoperiod of 6 h that was began and terminated by 5 or 15 min of light failed to rephase the rhythm.The shape of the curves for the rhythmic response of C. rubrum to the length of the dark period are sometimes suggestive of clocks operating on the principle of a tension-relaxation mechanism. Such a model allows for separate timing action of a rhythm and of P_fr disappearance over the early hours of darkness. Separate timing action does not, however, preclude an interaction between the rhythm and phytochrome in controlling flowering.Abbreviations FR far-red - P_fr far-red-absorbing form of phytochrome - P_r red-absorbing form of phytochrome - R red - BCJ photographic ruby-red irradiation A grant in aid of research from the National Research Council of Canada to B. G. Cumming is gratefully acknowledged.     

The involvement of phytochrome in controlling chlorophyll and 5-aminolevulinate formation in a gymnosperm seedling (Pinus sylvestris)

Chlorophyll a (Chl a) accumulation in the cotyledons of Scots pine seedlings (Pinus sylvestris L.) is much higher in the light than in darkness where it ceases 6 days after germination. When these darkgrown seedlings are treated with continuous white light (3,500 lx) a 3 h lag phase appears before Chl a accumulation is resumed. The lag phase can be eliminated by pretreating the seedlings with 7 h of weak red light (0.14 Wm^-2) or with 14 red light pulses separated by relatively short dark periods (<100 min). The effect of 15s red light pulses can be fully reversed by 1 min far-red light pulses. This reversibility is lost within 2 min. In addition, the amount of Chl a formed within 27 h of continuous red light is considerably reduced by the simultaneous application of far-red (RG 9) light. It is concluded that phytochrome (P_fr) is required not only for the elimination of the lagphase but also to maintain a high rate of Chl a accumulation in continuous light. Since accumulation of 5-aminolevulinate (ALA) responds in the same manner as Chl a accumulation to a red light pretreatment it is further concluded that ALA formation is the point where phytochrome regulates Chl biosynthesis in continuous light. No correlation has been found between ALA and Chl a formation in darkness. This indicates that in a darkgrown pine seedling ALA formation is not rate limiting for Chl a accumulation.Abbreviations Chl chlorophyll(ide) - PChl protochlorophyll(ide) - ALA 5-aminolevulinate - P_r the red absorbing form of phytochrome - P_fr the far-red absorbing form of phytochrome - P_tot total phytochrome ([P_r]+[P_fr])     

In-vitro binding of phytochrome to a particulate fraction: A function of light dose and steady-state P fr level

Summary The binding of phytochrome to a particulate fraction in extracts from hypocotyl hooks of etiolated Cucurbita pepo L. seedlings has been examined as a function of the light dose and P _fr level established in vitro. As the steady-state level of P _fr transiently established in the 500×g supernatant is increased, so the level of P _r subsequently pelletable at 20 000×g increases up to a saturation level. Increasing both the time and irradiance parameters of the light dose while holding the steady-state P _fr level constant, results similarly in increasing P _r pelletability. This agrees with results obtained previously with in-vivo irradiations of maize coleoptiles. Thus, like the in-vivo response, phytochrome binding in vitro appears to be a function of the total number of molecules converted to the P _fr form during the irradiation period.Abbreviations P _fr far-red-absorbing form of phytochrome - P _r red-absorbing form of phytochrome     

Phytochrome-mediated induction of phenylalanine ammonia-lyase in the cotyledons of tomato (Lycopersicon esculentum Mill.) plants

Phenylalanine ammonia-lyase (PAL; EC 4.3.1.5.) induction in cotyledons from 96-h dark-grown Lycopersicon esculentum Mill. was studied in response to continuous light and hourly light pulses (blue, red, far red). The increases of PAL promoted by blue and red pulses are reversed completely by immediately following 758 nm irradiations. The response to continuous red light could be substituted for by hourly 6-min red light pulses. The effect of continuous red treatments is mainly due to a multiple induction effect of phytochrome. In contrast to red light, hourly light pulses with far red and blue, light can only partially substitute for continuous irradiation. The continuous blue response could be due to a combination of a multiple induction response and of a high irradiance response of phytochrome. The continuous far red response, could represent a high irradiance response of phytochrome. Dichromatic irradiations indicate that phytochrome is the photoreceptor controlling the light response (PAL) in tomato seedlings.Abbreviations Norflurazon NF-4-chloro-5-(methylamino)-2-(,,,-trifluoro-m-tolyl)-3 (2H) pyridazinone - PAL phenylalanine ammonia-lyase - phytochrome photoequilibrium P_fr/P_tot - P_fr far-red absorbing form of phytochrome - P_r red absorbing form of phytochrome - P_tot total phytochrome: P_r+P_fr