phyA Mutant Research Articles

Interplay among photoreceptors determines the strategy of coping with excess light in tomato.

This study investigates photoreceptor's role in the adaption of photosynthetic apparatus to high light (HL) intensity by examining the response of tomato wild type (WT) (Solanum lycopersicum L. cv. Moneymaker) and tomato mutants (phyA, phyB1, phyB2, cry1) plants to HL. Our results showed a photoreceptor-dependent effect of HL on the maximum quantum yield of photosystem II (Fv/Fm) with phyB1 exhibiting a decrease, while phyB2 exhibiting an increase in Fv/Fm. HL resulted in an increase in the efficient quantum yield of photosystem II (ΦPSII) and a decrease in the non-photochemical quantum yields (ΦNPQ and ΦN0) solely in phyA. Under HL, phyA showed a significant decrease in the energy-dependent quenching component of NPQ (qE), while phyB2 mutants showed an increase in the state transition (qT) component. Furthermore, ΔΔFv/Fm revealed that PHYB1 compensates for the deficit of PHYA in phyA mutants. PHYA signaling likely emerges as the dominant effector of PHYB1 and PHYB2 signaling within the HL-induced signaling network. In addition, PHYB1 compensates for the role of CRY1 in regulating Fv/Fm in cry1 mutants. Overall, the results of this research provide valuable insights into the unique role of each photoreceptor and their interplay in balancing photon energy and photoprotection under HL condition.

Arogenate dehydratases: unique roles in light-directed development during the seed-to-seedling transition in Arabidopsis thaliana.

The seed-to-seedling transition is impacted by changes in nutrient availability and light profiles, but is still poorly understood. Phenylalanine affects early seedling development; thus, the roles of arogenate dehydratases (ADTs), which catalyze phenylalanine formation, were studied in germination and during the seed-to-seedling transition by exploring the impact of light conditions and specific hormone responses in adt mutants of Arabidopsis thaliana. ADT gene expression was assessed in distinct tissues and for light-quality dependence in seedlings for each of the six-member ADT gene family. Mutant adt seedlings were evaluated relative to wild type for germination, photomorphogenesis (blue, red, far red, white light, and dark conditions), anthocyanin accumulation, and plastid development-related phenotypes. ADT proteins are expressed in a light- and tissue-specific manner in transgenic seedlings. Among the analyzed adt mutants, adt3, adt5, and adt6 exhibit significant defects in germination, hypocotyl elongation, and root development responses during the seed-to-seedling transition. Interestingly, adt5 exhibits a light-dependent disruption in plastid development, similar to a phyA mutant. These data indicate interactions between photoreceptors, hormones, and regulation of phenylalanine pools in the process of seedling establishment. ADT5 and ADT6 may play important roles in coordinating hormone and light signals for normal early seedling development.

Arabidopsis FIN219/JAR1 interacts with phytochrome A under far-red light and jasmonates in regulating hypocotyl elongation via a functional demand manner.

Integration of light and phytohormones is essential for plant growth and development. FAR-RED INSENSITIVE 219 (FIN219)/JASMONATE RESISTANT 1 (JAR1) participates in phytochrome A (phyA)-mediated far-red (FR) light signaling in Arabidopsis and is a jasmonate (JA)-conjugating enzyme for the generation of an active JA-isoleucine. Accumulating evidence indicates that FR and JA signaling integrate with each other. However, the molecular mechanisms underlying their interaction remain largely unknown. Here, the phyA mutant was hypersensitive to JA. The double mutant fin219-2phyA-211 showed a synergistic effect on seedling development under FR light. Further evidence revealed that FIN219 and phyA antagonized with each other in a mutually functional demand to modulate hypocotyl elongation and expression of light- and JA-responsive genes. Moreover, FIN219 interacted with phyA under prolonged FR light, and MeJA could enhance their interaction with CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1) in the dark and FR light. FIN219 and phyA interaction occurred mainly in the cytoplasm, and they regulated their mutual subcellular localization under FR light. Surprisingly, the fin219-2 mutant abolished the formation of phyA nuclear bodies under FR light. Overall, these data identified a vital mechanism of phyA-FIN219-COP1 association in response to FR light, and MeJA may allow the photoactivated phyA to trigger photomorphogenic responses.

Regulation of Plant Photoresponses by Protein Kinase Activity of Phytochrome A.

Extensive research has been conducted for decades to elucidate the molecular and regulatory mechanisms for phytochrome-mediated light signaling in plants. As a result, tens of downstream signaling components that physically interact with phytochromes are identified, among which negative transcription factors for photomorphogenesis, PHYTOCHROME-INTERACTING FACTORs (PIFs), are well known to be regulated by phytochromes. In addition, phytochromes are also shown to inactivate an important E3 ligase complex consisting of CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) and SUPPRESSORs OF phyA-105 (SPAs). This inactivation induces the accumulation of positive transcription factors for plant photomorphogenesis, such as ELONGATED HYPOCOTYL 5 (HY5). Although many downstream components of phytochrome signaling have been studied thus far, it is not fully elucidated which intrinsic activity of phytochromes is necessary for the regulation of these components. It should be noted that phytochromes are autophosphorylating protein kinases. Recently, the protein kinase activity of phytochrome A (phyA) has shown to be important for its function in plant light signaling using Avena sativa phyA mutants with reduced or increased kinase activity. In this review, we highlight the function of phyA as a protein kinase to explain the regulation of plant photoresponses by phyA.

Stimulation of Tomato Drought Tolerance by PHYTOCHROME A and B1B2 Mutations.

Drought stress is a severe environmental issue that threatens agriculture at a large scale. PHYTOCHROMES (PHYs) are important photoreceptors in plants that control plant growth and development and are involved in plant stress response. The aim of this study was to identify the role of PHYs in the tomato cv. 'Moneymaker' under drought conditions. The tomato genome contains five PHYs, among which mutant lines in tomato PHYA and PHYB (B1 and B2) were used. Compared to the WT, phyA and phyB1B2 mutants exhibited drought tolerance and showed inhibition of electrolyte leakage and malondialdehyde accumulation, indicating decreased membrane damage in the leaves. Both phy mutants also inhibited oxidative damage by enhancing the expression of reactive oxygen species (ROS) scavenger genes, inhibiting hydrogen peroxide (H2O2) accumulation, and enhancing the percentage of antioxidant activities via DPPH test. Moreover, expression levels of several aquaporins were significantly higher in phyA and phyB1B2, and the relative water content (RWC) in leaves was higher than the RWC in the WT under drought stress, suggesting the enhancement of hydration status in the phy mutants. Therefore, inhibition of oxidative damage in phyA and phyB1B2 mutants may mitigate the harmful effects of drought by preventing membrane damage and conserving the plant hydrostatus.

Phytochrome A mediated modulation of photosynthesis, development and yield in rice (Oryza sativa L.) in fluctuating light environment

Phytochromes are sensory photoreceptors associated with the photomorphogenesis of plants. Rice has 3 phytochromes (phy); phyA, phyB and phyC. To understand the role of phyA under low light (LL) and normal light (NL) environments in plant growth, photosynthesis, biomass and yield, rice phytochrome A mutant (phyA) along with its wild-type (WT) genotype Akitakomachi were grown in field conditions. To induce LL (∼338 μmole photons m−2 s−1) both WT and phyA were shaded by the agro-shade net that reduced light intensity by ∼75%. The NL intensity (∼1355 μmole photons m−2 s−1) served as the experimental control. The plant height decreased by 8%−10% in phyA mutants both in NL and LL. Other developmental parameters, leaf area index, specific leaf weight, and spikelet fertility declined in the mutants in NL and to a larger extent in the LL. phyA had a lower electron transport rate (ETR), which increased its non-photochemical quenching (NPQ). The photosynthetic rate (PN), stomatal conductance(gs), transpiration rate (E), apparent quantum yield (AQE), maximum carboxylation efficiency (Vcmax) and mesophyll conductance (gm) significantly decreased in phyA than WT in LL. Furthermore, the expression of rice phytochrome-interacting factor-like protein (OsPIL1) was relatively downregulated in phyA under LL, resulting in a reduced total chlorophyll (Chl) and Chl b content and an increased Chl a/b ratio. Additionally, a relatively lower upregulation of Fv/Fm and expression of the isoforms of chlorophyll-a/b-proteins in phyA under LL suggests its inferior light-harvesting capacity compared to WT plants. Reduced PN resulted in a lowered overall carbon budget in phyA that hampered grain yield by 55% more than WT under LL. Our findings demonstrated the critical role of phytochrome A in orchestrating a series of shade-acclimation responses to salvage LL-induced stress and optimize the harvest index in rice.

Mutation of phytochrome B promotes resistance to sheath blight and saline-alkaline stress via increasing ammonium uptake in rice.

Phytochrome B (PhyB), a red-light receptor, plays important roles in diverse biological processes in plants; however, its function in NH4 + uptake and stress responses of plants is unclear. Here, we observed that mutation in indeterminate domain 10 (IDD10), which encodes a key transcription factor in NH4 + signaling, led to NH4 + -sensitive root growth in light but not in the dark. Genetic combinations of idd10 and phy mutants demonstrated that phyB, but not phyA or phyC, suppressed NH4 + -sensitive root growth of idd10. PhyB mutants and PhyB overexpressors (PhyB OXs) accumulated more and less NH4 + , respectively, compared with wild-type plants. Real time quantitative polymerase chain reaction (RT-qPCR) revealed that PhyB negatively regulated NH4 + -mediated induction of Ammonium transporter 1;2 (AMT1;2). AMT1 RNAi plants with suppressed AMT1;1, AMT1;2, and AMT1;3 expression exhibited shorter primary roots under NH4 + conditions. This suggested that NH4 + uptake might be positively associated with root growth. Further, PhyB interacted with and inhibited IDD10 and brassinazole-resistant 1 (BZR1). IDD10 interacted with BZR1 to activate AMT1;2. NH4 + uptake is known to promote resistance of rice (Oryza sativa) to sheath blight (ShB) and saline-alkaline stress. Inoculation of Rhizoctonia solani demonstrated that PhyB and IDD10 negatively regulated and AMT1 and BZR1 positively regulated resistance of rice to ShB. In addition, PhyB negatively regulated and IDD10 and AMT1 positively regulated resistance of rice to saline-alkaline stress. This suggested that PhyB-IDD10-AMT1;2 signaling regulates the saline-alkaline response, whereas the PhyB-BZR1-AMT1;2 pathway modulates ShB resistance. Collectively, these data prove that mutation in the PhyB gene enhances the resistance of rice to ShB and saline-alkaline stress by increasing NH4 + uptake.

Influence of phytochromes on microRNA expression, phenotype, and photosynthetic activity in A. thaliana phy mutants under light with different spectral composition

Light-induced changes in miRNAs, morphogenesis, and photosynthetic processes in phytochrome-deficient mutant plants grown under different light qualities were studied. miRNA activity in many processes is regulated by phytochromes and phytochrome-interacting factors (PIFs). The reduced content of photoreceptors in phytochrome mutants affects the PIF-microRNA interaction. In plants grown under red light (RL) and white light (WL), the phenotype of phyb mutant was distorted; however, under blue light (BL) conditions, the phyb phenotype was normalized. The photosynthetic rates of both the mutants and wild type were higher under BL than under RL and WL. The expression of most studied miRNAs increased in phyaphyb mutants under BL conditions, which is probably one of the reasons for the normalization of the phenotype, the increase in PSII activity, and the photosynthetic rate. MicroRNAs under BL can partially improve photosynthesis and phenotype of the mutants, which indicates the conjugation of the functioning of phytochromes in miRNA formation.

Functional Characterization of Tomato Phytochrome A and B1B2 Mutants in Response to Heat Stress.

Heat stress (HS) is a prevalent negative factor affecting plant growth and development, as it is predominant worldwide and threatens agriculture on a large scale. PHYTOCHROMES (PHYs) are photoreceptors that control plant growth and development, and the stress signaling response partially interferes with their activity. PHYA, B1, and B2 are the most well-known PHY types in tomatoes. Our study aimed to identify the role of tomato ‘Money Maker’ phyA and phyB1B2 mutants in stable and fluctuating high temperatures at different growth stages. In the seed germination and vegetative growth stages, the phy mutants were HS tolerant, while during the flowering stage the phy mutants revealed two opposing roles depending on the HS exposure period. The response of the phy mutants to HS during the fruiting stage showed similarity to WT. The most obvious stage that demonstrated phy mutants’ tolerance was the vegetative growth stage, in which a high degree of membrane stability and enhanced water preservation were achieved by the regulation of stomatal closure. In addition, both mutants upregulated the expression of heat-responsive genes related to heat tolerance. In addition to lower malondialdehyde accumulation, the phyA mutant enhanced proline levels. These results clarified the response of tomato phyA and phyB1B2 mutants to HS.

Effects of Green Light on Elongation Do Not Interact with Far-Red, Unless the Phytochrome Photostationary State (PSS) Changes in Tomato.

Simple SummaryThis paper focuses on the role of phytochromes (phys) in the interaction between green light and far-red light effects on “shade avoidance syndrome”. We grew wild type and phy mutants of tomato under a set of light conditions with different combinations of green, blue, red, and far-red light. Partial (20%) replacement of red/blue by green light in the absence of far-red light hardly affected the tomato plant morphology. However, when the spectrum contained far-red light, partially replacing red/blue by green light resulted in more elongation, which was associated with a lower phytochrome photostationary state (PSS) value. There was no effect of partial substitution of red/blue with green light when the PSS was kept constant. Thus, this study has revealed an interaction between green and far-red light effects on elongation unless PSS was kept constant. Green light was often a bit neglected in photobiology, but now an increasing number of researchers are realizing that green light deserves more attention. This study advances the understanding of light quality and plant growth and finding the optimal spectrum when growing plants under LED lighting in controlled environment agriculture.Green light (G) could trigger a “shade avoidance syndrome” (SAS) similarly to far-red light. We aimed to test the hypothesis that G interacts with far-red light to induce SAS, with this interaction mediated by phytochromes (phys). The tomato (Solanum lycopersicum cv. Moneymaker) wild-type (WT) and phyA, phyB1B2, and phyAB1B2 mutants were grown in a climate room with or without 30 µmol m−2 s−1 G on red/blue and red/blue/far-red backgrounds, maintaining the same photosynthetically active radiation (400–700 nm) of 150 µmol m−2 s−1 and red/blue ratio of 3. G hardly affected the dry mass accumulation or leaf area of WT, phyA, and phyB1B2 with or without far-red light. A lower phytochrome photostationary state (PSS) by adding far-red light significantly increased the total dry mass by enhancing the leaf area in WT plants but not in phy mutants. When the background light did not contain far-red light, partially replacing red/blue with G did not significantly affect stem elongation. However, when the background light contained far-red light, partially replacing red/blue with G enhanced elongation only when associated with a decrease in PSS, indicating that G interacts with far-red light on elongation only when the PSS changes.

Effect of high-intensity light and UV-B on photosynthetic activity and the expression of certain light-responsive genes in A. thaliana phyA and phyB mutants

The effects of high-intensity light (HIL, 4 and 24 h) and UV-B (1 h) on the net photosynthesis rate, activity of photosystem II (PSII), content of photosynthetic pigments, anthocyanin and UV-absorbing pigments as well as the expression of certain light-responsive genes (HY5,CAB1) chalcone synthase (CHS) and main antioxidants enzyme genes (APX1, GPX and GR) in the leaves of phyB and phyA mutant A. thaliana plants were studied. Both UV-B and 4 and 24 h HIL decreased the PSII maximum quantum yield (Fv/Fm), PSII performance index (PIABS), photosynthesis and respiration rates in plants. Moreover, all stress treatments increased the dissipation of the absorbed energy (DI0/RC) as well as the flux of absorbed energy per RC (ABS/RC). The maximal changes in photosynthesis and chlorophyll fluorescence parameters were observed in the phyB mutant. The WT and the phyA mutant showed similar responses. In addition, the phyB mutant exhibited decreases in the expression of genes encoding enzyme CHS, the transcription factor HY5 and the antioxidant enzymes APX1 and GPX. One of the possible mechanisms protecting the photosynthetic apparatus from light excess or UV radiation is the elevated content of various pigments that can act as antioxidants or optical filters. We assume that the greater decrease in photosynthetic activity in the phyB mutant under HIL and UV-B conditions was due to the decreased content of carotenoids and UV-absorbing pigments in this mutant.

Phytochromes control metabolic flux, and their action at the seedling stage determines adult plant biomass.

Phytochrome photoreceptors are known to regulate plastic growth responses to vegetation shade. However, recent reports also suggest an important role for phytochromes in carbon resource management, metabolism, and growth. Here, we use 13CO2 labelling patterns in multiallele phy mutants to investigate the role of phytochrome in the control of metabolic fluxes. We also combine quantitative data of 13C incorporation into protein and cell wall polymers, gas exchange measurements, and system modelling to investigate why biomass is decreased in adult multiallele phy mutants. Phytochrome influences the synthesis of stress metabolites such as raffinose and proline, and the accumulation of sugars, possibly through regulating vacuolar sugar transport. Remarkably, despite their modified metabolism and vastly altered architecture, growth rates in adult phy mutants resemble those of wild-type plants. Our results point to delayed seedling growth and smaller cotyledon size as the cause of the adult-stage phy mutant biomass defect. Our data signify a role for phytochrome in metabolic stress physiology and carbon partitioning, and illustrate that phytochrome action at the seedling stage sets the trajectory for adult biomass production.

Effect of phyB and phyC loss-of-function mutations on the wheat transcriptome under short and long day photoperiods

BackgroundPhotoperiod signals provide important cues by which plants regulate their growth and development in response to predictable seasonal changes. Phytochromes, a family of red and far-red light receptors, play critical roles in regulating flowering time in response to changing photoperiods. A previous study showed that loss-of-function mutations in either PHYB or PHYC result in large delays in heading time and in the differential regulation of a large number of genes in wheat plants grown in an inductive long day (LD) photoperiod.ResultsWe found that under non-inductive short-day (SD) photoperiods, phyB-null and phyC-null mutants were taller, had a reduced number of tillers, longer and wider leaves, and headed later than wild-type (WT) plants. The delay in heading between WT and phy mutants was greater in LD than in SD, confirming the importance of PHYB and PHYC in accelerating heading date in LDs. Both mutants flowered earlier in SD than LD, the inverse response to that of WT plants. In both SD and LD photoperiods, PHYB regulated more genes than PHYC. We identified subsets of differentially expressed and alternatively spliced genes that were specifically regulated by PHYB and PHYC in either SD or LD photoperiods, and a smaller set of genes that were regulated in both photoperiods. We found that photoperiod had a contrasting effect on transcript levels of the flowering promoting genes VRN-A1 and PPD-B1 in phyB and phyC mutants compared to the WT.ConclusionsOur study confirms the major role of both PHYB and PHYC in flowering promotion in LD conditions. Transcriptome characterization revealed an unexpected reversion of the wheat LD plants into SD plants in the phyB-null and phyC-null mutants and identified flowering genes showing significant interactions between phytochromes and photoperiod that may be involved in this phenomenon. Our RNA-seq data provides insight into light signaling pathways in inductive and non-inductive photoperiods and a set of candidate genes to dissect the underlying developmental regulatory networks in wheat.

Transcriptomic network analyses shed light on the regulation of cuticle development in maize leaves

Plant cuticles are composed of wax and cutin and evolved in the land plants as a hydrophobic boundary that reduces water loss from the plant epidermis. The expanding maize adult leaf displays a dynamic, proximodistal gradient of cuticle development, from the leaf base to the tip. Laser microdissection RNA Sequencing (LM-RNAseq) was performed along this proximodistal gradient, and complementary network analyses identified potential regulators of cuticle biosynthesis and deposition. A weighted gene coexpression network (WGCN) analysis suggested a previously undescribed function for PHYTOCHROME-mediated light signaling during the regulation of cuticular wax deposition. Genetic analyses reveal that phyB1 phyB2 double mutants of maize exhibit abnormal cuticle composition, supporting the predictions of our coexpression analysis. Reverse genetic analyses also show that phy mutants of the moss Physcomitrella patens exhibit abnormal cuticle composition, suggesting an ancestral role for PHYTOCHROME-mediated, light-stimulated regulation of cuticle development during plant evolution.

Nuclear-encoded sigma factor 6 (SIG6) is involved in the block of greening response in Arabidopsis thaliana.

Light is critical in the ability of plants to accumulate chlorophyll. When exposed to far-red (FR) light and then grown in white light in the absence of sucrose, wild-type seedlings fail to green in a response known as the FR block of greening (BOG). This response is controlled by phytochrome A through repression of protochlorophyllide reductase-encoding (POR) genes by FR light coupled with irreversible plastid damage. Sigma (SIG) factors are nuclear-encoded proteins that contribute to plant greening and plastid development through regulating gene transcription in chloroplasts and impacting retrograde signaling from the plastid to nucleus. SIGs are regulated by phytochromes, and the expression of some SIG factors is reduced in phytochrome mutant lines, includingphyA. Given the association of phyA with the FR BOG and its regulation of SIG factors, we investigated the potential regulatory role of SIG factors in the FR BOG response. We examined FR BOG responses in sig mutants, phytochrome-deficient lines, and mutant lines for several phy-associated factors. We quantified chlorophyll levels and examined expression of key BOG-associated genes. Among six sig mutants, onlythe sig6mutant significantly accumulated chlorophyll after FR BOG treatment, similar tothe phyA mutant. SIG6 appears to control protochlorophyllide accumulation by contributing to the regulation of tetrapyrrole biosynthesis associated with glutamyl-tRNA reductase (HEMA1) function, select phytochrome-interacting factor genes (PIF4 and PIF6), and PENTA1, which regulates PORA mRNA translation after FR exposure. Regulation of SIG6 plays a significant role in plant responses to FR exposure during the BOG response.

The Impacts of Varying Types of Light on the Growth of Five Arabidopsis Varieties

Arabidopsis is known as the “fruit fly of plants.” It is small and easy to grow, has a short life cycle, and has a small, easy-to-manipulate genome. Using Arabidopsis, we tested the effects of varied light conditions on the plant growth of mutants with dysfunctional light pathways. We tested five different strains: wild type, a phytochrome A mutant (phyA), a phytochrome B mutant (phyB), a phyA/phyB double mutant, and a DET1-1 mutant. With these mutants, we investigated how varied wavelengths and exposure of light affect the growth of the mutants. We found that the phyA mutant, the phyB mutant, and the double mutant all grew well in red light, with high germination rates and the largest average plant size. The phyB mutant grew the best under blue light, with the highest germination rate and the second largest average plant size. Under natural light, every strain grew relatively well, with high germination rates and consistent sizes. Although the DET1-1 mutant had a lower average size compared to the phyA and phyB mutations, it had the highest germination percentage, making it the most successful under no-light conditions.

Phytochrome A Negatively Regulates the Shade Avoidance Response by Increasing Auxin/Indole Acidic Acid Protein Stability

The reduction in the red to far-red light ratio (R/FR) and photosynthetically active radiation caused by dense planting initiates shade avoidance responses (SARs) to help plants compete against their neighbors. However, deep shade attenuates shade-induced stem elongation to suppress excessive reversion toward skotomorphogenic development, in which photoreceptor phytochrome A (PHYA) has been known to play the major role. However, the molecular mechanism underlying PHYA function in deep shade is poorly understood. Here, we report that shade-accumulated PHYA can release auxin/indole-3-acetic acid (AUX/IAA), suppressors in the auxin signaling pathway, from SCFTIR1, an auxin receptor, to weaken auxin signaling and negatively regulate shade response. Corroborating this, phyA mutants display an enhanced auxin response to deep shade and auxin treatment. Specifically, PHYA competes with TIR1 by directly binding and stabilizing AUX/IAA. Our findings illustrate a mechanistic model of how plants sense different shade levels to fine-tune auxin signaling and generate appropriate SAR.

Biological activity and dimerization state of modified phytochrome A proteins.

To assess potential physical interactions of type I phyA with the type II phyB-phyE phytochromes in vivo, transgenes expressing fusion gene forms of phyA were introduced into the Arabidopsis phyA mutant background. When a single c-Myc (myc) epitope is added to either the N- or C-terminus of phyA, the constructs completely complement phyA mutant phenotypes. However, addition of larger tags, such as six consecutive myc epitopes or the yellow fluorescent protein sequence, result in fusion proteins that show reduced activity. All the tagged phyA proteins migrate as dimers on native gels and co-immunoprecipitation reveals no binding interaction of phyA to any of the type II phys in the dark or under continuous far-red light. Dimers of the phyA 1–615 amino acid N-terminal photosensory domain (NphyA), generated in vivo with a yeast GAL4 dimerization domain and attached to a constitutive nuclear localization sequence, are expressed at a low level and, although they cause a cop phenotype in darkness and mediate a very low fluence response to pulses of FR, have no activity under continuous FR. It is concluded that type I phyA in its Pr form is present in plants predominantly or exclusively as a homodimer and does not stably interact with type II phys in a dimer-to-dimer manner. In addition, its activity in mediating response to continuous FR is sensitive to modification of its N- or C-terminus.

Improved thermostability and enzyme activity of a recombinant phyA mutant phytase from Aspergillus niger N25 by directed evolution and site-directed mutagenesis.

We previously constructed three recombinant phyA mutant strains (PP-NPm-8, PP-NPep-6A and I44E/T252R-PhyA), showing improved catalytic efficiency or thermostability of Aspergillus niger N25 phytase, by error-prone PCR or site-directed mutagenesis. In this study, directed evolution and site-directed mutagenesis were further applied to improve the modified phytase properties. After one-round error-prone PCR for phytase gene of PP-NPep-6A, a single transformant, T195L/Q368E/F376Y, was obtained with the significant improvements in catalytic efficiency and thermostability. The phytase gene of T195L/Q368E/F376Y, combined with the previous mutant phytase genes of PP-NPep-6A, PP-NPm-8 and I44E/T252R-PhyA, was then sequentially modified by DNA shuffling. Three genetically engineered strains with desirable properties were then obtained, namedQ172R, Q172R/K432R andQ368E/K432R. Among them, Q172R/K432R showed the highest thermostability with the longest half-life and the greatest remaining phytase activity after heat treatment, while Q368E/K432R showed the highest catalytic activity. Five substitutions (Q172R, T195L, Q368E, F376Y, K432R) identified from random mutagenesis were added sequentially to the phytase gene of PP-NPep-6A to investigate how the mutant sites influence the properties of phytase. Characterization and structural analysis demonstrated that these mutations could produce cumulative or synergistic improvements in thermostability or catalytic efficiency of phytase.

Phytochromes are key regulators of abiotic stress responses in tomato

Phytochromes are the best characterized and most frequently studied plant photoreceptors. A plethora of studies have revealed important roles for phytochromes in plant development, and more recently, evidence indicates that these photoreceptors also modulate responses to a multitude of abiotic and biotic stresses. Thus, the present work aimed to investigate whether tomato phytochromes phyA, phyB1 and phyB2 are involved with responses to low water potential (polyethylene glycol 6000 at Ψw of −0.3MPa), high salinity (100mM NaCl), cadmium contamination (65mM CdCl2), high temperature (42°C for six hours during three days) and ultraviolet B radiation (UV-B − 280–320nm for eight hours during three days) stresses. For this purpose, seedlings of tomato mutants impacted by phytochrome A (fri), phytochrome B1 (tri) and phytochrome B2 (phyB2) were subjected to abiotic stresses and evaluated for their growth, pigment and osmoprotectant accumulation and lipid peroxidation. Under the conditions of this study, the results did not shown large variations of phyA mutant when compared to the wild genotype. However, the tomato phytochromes B1 and B2 mainly act as negative regulators of growth, pigment maintenance and osmoprotectant accumulation during responses to the different abiotic stresses.