The 14-3-3 Proteins mu and upsilon influence transition to flowering and early phytochrome response

14-3-3 proteins regulate a diverse set of biological responses but developmental phenotypes associated with 14-3-3 mutations have not been described in plants. Here, physiological and biochemical tests demonstrate interactions between 14-3-3s and the well-established mechanisms that govern light sensing and photoperiodic flowering control. Plants featuring homozygous disruption of 14-3-3 isoforms upsilon and mu display defects in light sensing and/or response. Mutant plants flower late and exhibit long hypocotyls under red light, with little effect under blue or far-red light. The long hypocotyl phenotype is consistent with a role for 14-3-3 upsilon and mu in phytochrome B signaling. Yeast two-hybrid and coimmunoprecipitation assays indicate that 14-3-3 upsilon and mu proteins physically interact with CONSTANS, a central regulator of the photoperiod pathway. Together, these data indicate a potential role for specific 14-3-3 isoforms in affecting photoperiodic flowering via interaction with CONSTANS, possibly as integrators of light signals sensed through the phytochrome system.     

Phytochromes confer the photoperiodic control of flowering in rice (a short-day plant)

The photoperiodic sensitivity 5 (se5) mutant of rice, a short-day plant, has a very early flowering phenotype and is completely deficient in photoperiodic response. We have cloned the SE5 gene by candidate cloning and demonstrated that it encodes a putative heme oxygenase. Lack of responses of coleoptile elongation by light pulses and photoreversible phytochromes in crude extracts of se5 indicate that SE5 may function in phytochrome chromophore biosynthesis. Ectopic expression of SE5 cDNA by the CaMV 35S promoter restored the photoperiodic response in the se5 mutant. Our results indicate that phytochromes confer the photoperiodic control of flowering in rice. Comparison of se5 with hy1, a counterpart mutant of Arabidopsis, suggests distinct roles of phytochromes in the photoperiodic control of flowering in these two species.     

Ectopic expression of oat phytochrome A in hybrid aspen changes critical daylength for growth and prevents cold acclimatization

Survival of temperate-zone tree species under the normal summer-winter cycle is dependent on proper timing of apical growth cessation and cold acclimatization. This timing is primarily based on the perception of daylength, and through evolution many tree species have developed photoperiodic ecotypes which are closely adapted to the local light conditions. The longest photoperiod inducing growth cessation, the critical photoperiod, is inherited as a quantitative character. The phytochrome pigment family is the probable receptor of daylength, but the exact role of phytochrome and the physiological basis for the different responses between photoperiodic ecotypes are not known. This report shows for the first time that over-expression of the oat phytochrome A gene ( PHYA ) in a tree significantly changes the critical daylength and effectively prevents cold acclimatization. While the critical daylength for elongation growth in the wild-type of hybrid aspen ( Populus tremula × tremuloides ) was approximately 15 h, transgenic lines with a strong expression of the oat PHYA gene did not stop growing even under a photoperiod of 6 h. Quantitative analysis of gibberellins (GA) as well as indole-3-acetic acid (IAA) revealed that levels of these were not down-regulated under short days in the transgenic plants expressing high levels of oat PHYA , as in the wild-type. These results indicate that photoperiodic responses in trees might be regulated by the amount of PHYA gene expressed in the plants, and that the amount of phytochrome A (phyA) affects the metabolism of GAs and IAA.     

14-3-3 isoforms participate in red light signaling and photoperiodic flowering

Members of the 14-3-3 family of proteins participate in signal transduction by modulating flux through various pathways. Potential subfunctionalization within this family has produced a suite of related proteins with diverse client interactions and discrete localization. The associated study assesses the biological roles of two specific 14-3-3 isoforms, using genetic, biochemical and physiological assays to ascertain potential nodes of interaction. Arabidopsis T-DNA insertion mutants representing the ν and μ isoforms exhibited a short, yet clear delay in flowering time on long days. Tests of hypocotyl growth inhibition under narrow bandwidth light indicated a hyposensitivity to red light, while responses to blue and far-red light were normal. These physiological tests suggest a mechanistic link between 14-3-3 proteins, red light sensing, and the pathways that control photoperiodic flowering. The precise entry point into the pathway was assessed using yeast two hybrid assays targeted against specific proteins active in the circadian oscillator, light transduction and photoperiodic flowering. Yeast two hybrid interaction was observed with CONSTANS (CO), and then confirmed with coimmunoprecipitation. Functional interaction with phyB leading to defects in flowering time and direct interaction with CONSTANS circumstantially places these specific 14-3-3 isoforms into the pathway that regulates the transition between vegetative and floral development.Key words: isoform specificity, protein interaction, phosphorylation, signaling     

植物光信号途径重要新调控因子TZP的研究进展

TZP(TANDEM ZINC-FINGER/PLUS3)是近年来鉴定到的一个光信号转导途径新组分,在光介导的植物生长发育过程中发挥重要调控作用。TZP不仅负调控蓝光信号途径,参与光敏色素B(phyB)介导的开花调控过程,还参与调控phyA在体内的蛋白质磷酸化。对TZP生化活性和作用机制的深入研究,不仅有助于进一步完善光信号调控网络,也可为设计和培育具有耐密理想株型及高光效作物新品种提供理论依据。该文系统总结了TZP在植物光信号途径中发挥的重要调控作用,并提出未来TZP功能研究的重要问题。     

Mutations in the gene for the red/far-red light receptor phytochrome B alter cell elongation and physiological responses throughout Arabidopsis development.

Phytochromes are a family of plant photoreceptors that mediate physiological and developmental responses to changes in red and far-red light conditions. In Arabidopsis, there are genes for at least five phytochrome proteins. These photoreceptors control such responses as germination, stem elongation, flowering, gene expression, and chloroplast and leaf development. However, it is not known which red light responses are controlled by which phytochrome species, or whether the different phytochromes have overlapping functions. We report here that previously described hy3 mutants have mutations in the gene coding for phytochrome B (PhyB). These are the first mutations shown to lie in a plant photoreceptor gene. A number of tissues are abnormally elongated in the hy3(phyB) mutants, including hypocotyls, stems, petioles, and root hairs. In addition, the mutants flower earlier than the wild type, and they accumulate less chlorophyll. PhyB thus controls Arabidopsis development at numerous stages and in multiple tissues.     

Bottom-up Assembly of the Phytochrome Network

Plants have developed sophisticated systems to monitor and rapidly acclimate to environmental fluctuations. Light is an essential source of environmental information throughout the plant’s life cycle. The model plant Arabidopsis thaliana possesses five phytochromes (phyA-phyE) with important roles in germination, seedling establishment, shade avoidance, and flowering. However, our understanding of the phytochrome signaling network is incomplete, and little is known about the individual roles of phytochromes and how they function cooperatively to mediate light responses. Here, we used a bottom-up approach to study the phytochrome network. We added each of the five phytochromes to a phytochrome-less background to study their individual roles and then added the phytochromes by pairs to study their interactions. By analyzing the 16 resulting genotypes, we revealed unique roles for each phytochrome and identified novel phytochrome interactions that regulate germination and the onset of flowering. Furthermore, we found that ambient temperature has both phytochrome-dependent and -independent effects, suggesting that multiple pathways integrate temperature and light signaling. Surprisingly, none of the phytochromes alone conferred a photoperiodic response. Although phyE and phyB were the strongest repressors of flowering, both phyB and phyC were needed to confer a flowering response to photoperiod. Thus, a specific combination of phytochromes is required to detect changes in photoperiod, whereas single phytochromes are sufficient to respond to light quality, indicating how phytochromes signal different light cues.     

Mutational analyses of light-controlled seedling development in Arabidopsis

Arabidopsis mutants with decreased responses to light and mutants showing light responses in the dark have both been characterized. Some of the former mutants lack specific photoreceptors, such as the red/far-red light receptor phytochrome A, phychrome B, or a putative blue light receptor, HY4. These have allowed the assessment of physiological functions of these photoreceptors. The mutants with light responses in the dark include some, such as det1 and cop1, that appear to identify light signal transduction components, and others, such as fus6, that may be less directly related to normal control of light responses. Double mutant studies suggest how the different gene products might interact.     

Induction of diapause and seasonal morphs in butterflies and other insects: knowns,unknowns and the challenge of integration

The ‘choice’ of whether to enter diapause or to develop directly has profound effects on the life histories of insects, and may thus have cascading consequences such as seasonal morphs and other less obvious forms of seasonal plasticity. Present knowledge of the control of diapause and seasonal morphs at the physiological and molecular levels is briefly reviewed. Examples, mainly derived from personal research (primarily on butterflies), are given as a starting point with the aim of outlining areas of research that are still poorly understood. These include: the role of the direction of change in photoperiod; the role of factors such as temperature and diet in modifying the photoperiodic responses; and the role of sex, parental effects and sex linkage on photoperiodic control. More generally, there is still a limited understanding of how external cues and physiological pathways regulating various traits are interconnected via gene action to form a co‐adapted complete phenotype that is adaptive in the wild despite environmental fluctuation and change.     

Degradation of phytochrome A and the high irradiance response in Arabidopsis: a kinetic analysis

The ability to respond to far‐red‐rich light is essential for seedlings germinating below dense canopies. Physiological and genetic studies have demonstrated that phytochrome A is the only photoreceptor mediating responses to far‐red light. However, all phytochromes including phytochrome A are believed to be activated by red light and to be inactivated by far‐red light. To address the fundamental question of why phytochrome A has its highest physiological activity at presumably inactivating wavelengths, we analysed light‐induced degradation of phytochrome A in Arabidopsis. Rate constants were obtained for all reaction events in a two‐step model of degradation. Based on biochemical data, the model includes a tagging mechanism preceding degradation. The parameterized model describes Pr accumulation, wavelength dependencies of degradation kinetics and steady‐state levels as well as Pfr‐induced Pr degradation. Subsequently, experimentally derived fluence rate response curves, action spectrum and response curves to dichromatic irradiation were compared to simulations based on the model of degradation. Two kinetically defined phytochrome subspecies, untagged Pfr and tagged Pr, have steady‐state levels closely matching the physiological response curves. Therefore, sensing of far‐red light by phytochrome A can be quantitatively explained based exclusively on regulated protein degradation.     

Photoperiodic control of cytokinin transport and metabolism in Chenopodium rubrum

Cytokinin (CK) levels in the short-day plant Chenopodium rubrum L. are known to fluctuate diurnally. The aim of this work was to investigate if the diurnal changes are brought about by changes in transport and/or metabolism of CKs. The effect of photo-period on cytokinin transport was studied by analysing CK concentrations in root, leaf and apical exudates, respectively, under constant light (CL), a 12-h photoperiod (DL) inductive for flowering, DL in which darkness was interrupted at the end of hour 6 by 15 min red light (R), or by 15 min R followed by 30 min far-red irradiation (R/FR). The concentrations of cytokinins (zeatin, zeatin riboside, isopentenyladenine, isopen-tenyladenosine) in all three types of exudates were significantly higher in the first 12-h period after the end of 12 h darkness than in CL. The R break almost fully negated the effect of darkness and its effect was reversed by FR, showing the involvement of phytochrome in the regulation of CK transport. In the next 12-h interval, i.e. 12–24 h after the end of darkness, the CK level remained high in the leaf exudate only, but to a much lower extent than in the previous 12 h. The highest CK concentration (increase by 108%) was observed in apical exudates during inductive darkness. A comparison of the CKs present in the individual exudates indicates that those arriving at the apical part are derived mostly from leaves with varying contributions by the xylem. The metabolism of applied [³H]-zeatin riboside (ZR) was studied using HPLC separation of the metabolites. Metabolism was found to be very rapid and different glucosides, adenine and adenosine were the main metabolites after 12 h incubation with labelled ZR in all regimes tested. The only metabolite that seems to be under photoperiodic control is ZR-5′-monophosphate. It is as yet not clear if photoperiod controls the phosphorylation or dephosphorylation reaction. The activity of the main cytokinin degradative enzyme, cytokinin oxidase, did not change during the photoperiodic regimes tested.     

Phytochromes are Pr-ipatetic kinases.

A series of new studies reveal how the red/far-red light photoreceptors called phytochromes act. Phytochrome A and phytochrome B each move to the nucleus when activated by light, and phytochrome A is a kinase. Phytochrome-interacting proteins provide candidate signal transduction components and a recent physiological study suggests how phyA may mediate responses to far-red light. Regulation of phytochrome nuclear localization and kinase activities creates multiple phytochrome species, which may each have different regulatory activities.     

Phytochrome B and the Regulation of Circadian Ethylene Production in Sorghum

The sorghum (Sorghum bicolor L. Moench) cultivar 58M, which contains the null mutant phytochrome B gene, shows reduced photoperiodic sensitivity and exhibits a shade-avoidance phenotype. Ethylene production by seedlings of wild-type and phytochrome B mutant cultivars was monitored every 3 h, and both cultivars were found to produce ethylene in a circadian rhythm, with peak production occurring during the day. The phytochrome B mutant produces rhythmic peaks of ethylene with approximately 10 times the amplitude of the wild-type counterpart with the same period and diurnal timing. The source of the mutant's additional ethylene is the shoot. The diurnal rhythm can be produced with either light or temperature cycles; however, both light and temperature cycles are required for circadian entrainment. The temperature signal overrides the light signal in the production of diurnal rhythms, because seedlings grown under thermoperiods reversed with the photoperiod produced ethylene peaks during the warm nights. To examine the effect of extreme shading on ethylene production, seedlings were grown under dim, far-red-enriched light. This treatment duplicated the phytochrome B mutant's shade-avoidance phenotype in the wild type and caused the wild type to produce ethylene peaks similar to those observed in the mutant. The results confirm that phytochrome B is not required for proper function of circadian timing, but it may be involved in modulating physiological rhythms driven by the biological clock oscillator.     

Overexpression of the Heterotrimeric G-Protein α-Subunit Enhances Phytochrome-Mediated Inhibition of Hypocotyl Elongation in Arabidopsis

Plant heterotrimeric G-proteins have been implicated in a number of signaling processes. However, most of these studies are based on biochemical or pharmacological approaches. To examine the role of heterotrimeric G-proteins in plant development, we generated transgenic Arabidopsis expressing the Galpha subunit of the heterotrimeric G-protein under the control of a glucocorticoid-inducible promoter. With the conditional overexpression of either the wild type or a constitutively active version of Arabidopsis Galpha, transgenic seedlings exhibited a hypersensitive response to light. This enhanced light sensitivity was more exaggerated in a relatively lower intensity of light and was observed in white light as well as far-red, red, and blue light conditions. The enhanced responses in far-red and red light required functional phytochrome A and phytochrome B, respectively. Furthermore, the response to far-red light depended on functional FHY1 but not on FIN219 and FHY3. This dependence on FHY1 indicates that the Arabidopsis Galpha protein may act only on a discrete branch of the phytochrome A signaling pathway. Thus, our results support the involvement of a heterotrimeric G-protein in the light regulation of Arabidopsis seedling development.     

Transposing phytochrome into the nucleus

To control many physiological responses, phytochromes directly modulate gene expression. A key regulatory event in this signal transduction pathway is the light-controlled translocation of the photoreceptor from the cytoplasm into the nucleus. Recent publications are beginning to shed light on the molecular mechanisms underlying this central control point. Interestingly, there is a specific mechanism for phytochrome A (phyA) nuclear accumulation. The dedicated phyA nuclear import pathway might be important for the distinct photosensory specificity of this atypical phytochrome. Recent studies in the field also provide a starting point for investigating how the different subcellular pools of phytochrome can control distinct responses to light.     

Phytochrome E Controls Light-Induced Germination of Arabidopsis

Germination of Arabidopsis seeds is light dependent and under phytochrome control. Previously, phytochromes A and B and at least one additional, unspecified phytochrome were shown to be involved in this process. Here, we used a set of photoreceptor mutants to test whether phytochrome D and/or phytochrome E can control germination of Arabidopsis. The results show that only phytochromes B and E, but not phytochrome D, participate directly in red/far-red light (FR)-reversible germination. Unlike phytochromes B and D, phytochrome E did not inhibit phytochrome A-mediated germination. Surprisingly, phytochrome E was required for germination of Arabidopsis seeds in continuous FR. However, inhibition of hypocotyl elongation by FR, induction of cotyledon unfolding, and induction of agravitropic growth were not affected by loss of phytochrome E. Therefore, phytochrome E is not required per se for phytochrome A-mediated very low fluence responses and the high irradiance response. Immunoblotting revealed that the need of phytochrome E for germination in FR was not caused by altered phytochrome A levels. These results uncover a novel role of phytochrome E in plant development and demonstrate the considerable functional diversification of the closely related phytochromes B, D, and E.     

Overexpressed phytochrome C has similar photosensory specificity to phytochrome B but a distinctive capacity to enhance primary leaf expansion

Phytochrome C (phyC) is a low-abundance member of the five-membered phytochrome family of photoreceptors in Arabidopsis. Towards developing an understanding of the photosensory and physiological functions of phyC, transgenic Arabidopsis plants were generated that overexpress cDNA-encoded phyC and seedling responses to continuous white, red, or far-red light (Wc, Rc or FRc, respectively) were examined. Transgenic seedlings overexpressing phyC displayed enhanced inhibition of hypocotyl elongation in Rc, but were unchanged in responsiveness to FRc relative to wild-type. These data indicate that phyC has photosensory specificity that is similar to that of phyB and thus distinct from that of phyA. phyC overexpressors with levels only 3 to 4 times the level of endogenous phyC exhibited enhanced primary leaf expansion in Wc. This is in contrast to phyA or phyB overexpressors which respectively have levels that are 500-and 100-fold that of overexpressed phyC but showed no enhancement of primary leaf expansion. Therefore, phyC may have some physiological roles that are different to those of phyA and phyB in the control of seedling responses to light signals.     

不同光敏色素分子的特异感光性

简要综述两种光敏色素（ＰｈｙＡ、ＰｈｙＢ） 的分子特性、感光性及作用模式等方面的研究进展。光敏色素是一种调节植物中许多光反应的色素蛋白复合体。不同光敏色素分子具有特异的感光性。ＰｈｙＡ负责‘甚低辐照反应’和远红光‘高辐照反应’; 而ＰｈｙＢ则调节‘低辐照反应’及红光‘高辐照反应’。另外, 讨论了ＰｈｙＡ和ＰｈｙＢ在光周期感受中的作用。     

The phytochrome photoreceptor in the green alga Mesotaenium caldariorum: implication for a conserved mechanism of phytochrome action

Chloroplast movement in the unicellular green alga Mesotaenium caldariorum is one of the earliest documented photomorphogenetic responses in plants. Photobiological studies have established that this response is under the control of phytochrome, whose rigid association with the plasma membrane and/or cytoskeleton enables the algal cells to orientate the chloroplast in response to the direction and intensity of light from the environment. While many of the key components of the algal phytochrome signalling pathway have been elucidated (i.e. Ca²⁺, calmodulin, actin and myosin), the primary biochemical mechanism of algal phytochrome action is unknown. To begin to address this important question, phytochrome and its corresponding genes have been isolated and characterized in this alga. These studies reveal that Mesotaenium cells contain a single type of phytochrome which is encoded by a small family of highly related genes. On the basis of its biochemical properties, primary structure and ability to interfere with the photoregulatory activity of phytochrome in transgenic plant seedlings, it appears likely that the primary mechanism of phytochrome action has been conserved throughout its evolution.     

Phytochrome B mediates the photoperiodic control of tuber formation in potato

To determine whether phytochrome B is involved in the response of potato plants to photoperiod, a potato PHYB cDNA fragment was inserted in the antisense orientation behind the 35S CaMV promoter in Bin19 and this construct was transformed into Solanum tuberosum ssp. andigena plants which normally require short days for tuberization. Two independent transformants were obtained that had much lower levels of PHYB mRNA and protein, and which exhibited phenotypes characteristic of phyB mutants, for example, elongated stems and decreased chlorophyll content. The level of phyA, and of several phytochrome A-controlled responses, was unaffected in these plants. The photoperiodic control of tuberization in these antisense PHYB plants was abolished, the plants tuberizing in short day, long day, or short day plus night break conditions. This result shows that phytochrome B is required for the photoperiodic control of tuberization in potato ( Solanum tuberosum ssp. andigena ) and that it regulates this developmental process by preventing tuber formation in non-inductive photoperiods rather than by promoting tuberization in inductive photoperiods.