Phot2 Mutants Research Articles

Both LOV1 and LOV2 domains of phototropin2 function as the photosensory domain for hypocotyl phototropic responses in Arabidopsis thaliana (Brassicaceae)

Phototropins (phot) are blue light receptor proteins that mediate phototropism and control photomovement responses, such as chloroplast photorelocation movement and stomatal opening. Arabidopsis thaliana has two phototropins, phot1 and phot2. Although both phot1 and phot2 redundantly mediate photomovement responses, phot2 uniquely regulates phototropism and the chloroplast avoidance response under high-intensity blue light. However, compared to that of phot1, the mechanistic basis of phot2 function is poorly understood, and in particular, the importance of the LOV2 domain in phot2 function has not been clearly demonstrated. Indeed, photocycle-deficient LOV2 transgenic lines expressing phot2 in a phot1phot2 mutant background retained phototropism, although with less sensitivity than wild-type plants. We isolated 11 alleles of phot2 mutants and determined the molecular lesion in each allele. We analyzed hypocotyl phototropism, chloroplast photorelocation movement, and leaf flattening in the phot2 mutant and the respective phot1phot2 double mutant plants. We demonstrated that unlike the phot2 null mutant, the phot2-10 mutant, which has the defective phot2 LOV2 domain, retained the phototropic response and had unusual chloroplast movement. Mutants phot2-2 and phot2-6, which have a missense mutation in the kinase activation loop of phot2, had the phot2-null mutant phenotype. Furthermore, we convincingly demonstrated that the commonly used phot2-1 mutant allele is a phot2-null mutant. The analyses of the multiple phot2 mutant alleles provided strong evidence for the importance of both LOV domains and the kinase activation loop of phot2 in phototropism and other phot-dependent responses and also demonstrated that phot2-1 allele is a null mutant.

Phototropin 1 and cryptochrome action in response to green light in combination with other wavelengths

Genetic studies have shown the effects of various photoreceptors on early photomorphogenic processes, defining the precise time course of red (RL), far-red (FrL) and blue light (BL) action. In this study, the effect of green wavebands in conjunction with these responses is examined. Longer-term (end point; 24-96 h) analysis of hypocotyl elongation in enriched green environments shows an increase in growth compared to seedlings under blue, red or both together. The effect was only observed at lower fluence rates (<10 μmol/m² s). Genetic analyses demonstrate that cryptochromes are required for this GL effect, consistent with earlier findings, and that the phy receptors have no influence. However, analysis of early (minutes to hours) stem growth kinetics indicates that GL cannot reverse the cryptochrome-mediated BL effect during early stem growth inhibition, and instead acts additively with BL to drive cryptochrome-mediated inhibition. Green light (GL) treatments antagonize RL and FrL-mediated hypocotyl inhibition. The GL opposition of RL responses persists in phyA, phyB, cry1cry2 and phot2 mutants. The response requires phot1 and NPH3, suggesting that this is not a GL response, but instead a response to extremely low-fluence rate BL. Tests with dim BL (<0.1 μmol/m² s) confirm a previously uncharacterized phot1-dependent promotion of stem growth, opposing the effects of RL. These findings demonstrate how enriched green environments may adjust RL and BL photomorphogenic responses through both the crys and phot1 receptors, and define a new role for phot1 in stem growth promotion.

The Role of a 14-3-3 Protein in Stomatal Opening Mediated by PHOT2 in Arabidopsis

The 14-3-3 λ isoform is required for normal stomatal opening mediated by PHOT2 in Arabidopsis thaliana. Arabidopsis phototropin2 (PHOT2) interacts with the λ-isoform 14-3-3 protein both in yeast two-hybrid screening and in an in vitro pull-down assay. Further yeast two-hybrid analysis also showed that the PHOT2 C-terminal kinase domain was required for the interaction. Site-directed mutagenesis indicated that PHOT2 Ser-747 is essential for the yeast interaction. Phenotypic characterization of a loss-of-function 14-3-3 λ mutant in a phot1 mutant background showed that the 14-3-3 λ protein was necessary for normal PHOT2-mediated blue light-induced stomatal opening. PHOT2 Ser-747 was necessary for complementation of the blue light-activated stomatal response in a phot1 phot2 double mutant. The 14-3-3 λ mutant in the phot1 mutant background allowed normal phototropism and normal chloroplast accumulation and avoidance responses. It also showed normal stomatal opening mediated by PHOT1 in a phot2 mutant background. The 14-3-3 κ mutant had no effect on stomatal opening in response to blue light. Although the 14-3-3 λ mutant had no chloroplast movement phenotype, the 14-3-3 κ mutation caused a weaker avoidance response at an intermediate blue light intensity by altering the balance between the avoidance and accumulation responses. The results highlight the strict specificity of phototropin-mediated signal transduction pathways.

The Signal Transducer NPH3 Integrates the Phototropin1 Photosensor with PIN2-Based Polar Auxin Transport in Arabidopsis Root Phototropism

Under blue light (BL) illumination, Arabidopsis thaliana roots grow away from the light source, showing a negative phototropic response. However, the mechanism of root phototropism is still unclear. Using a noninvasive microelectrode system, we showed that the BL sensor phototropin1 (phot1), the signal transducer NONPHOTOTROPIC HYPOCOTYL3 (NPH3), and the auxin efflux transporter PIN2 were essential for BL-induced auxin flux in the root apex transition zone. We also found that PIN2-green fluorescent protein (GFP) localized to vacuole-like compartments (VLCs) in dark-grown root epidermal and cortical cells, and phot1/NPH3 mediated a BL-initiated pathway that caused PIN2 redistribution to the plasma membrane. When dark-grown roots were exposed to brefeldin A (BFA), PIN2-GFP remained in VLCs in darkness, and BL caused PIN2-GFP disappearance from VLCs and induced PIN2-GFP-FM4-64 colocalization within enlarged compartments. In the nph3 mutant, both dark and BL BFA treatments caused the disappearance of PIN2-GFP from VLCs. However, in the phot1 mutant, PIN2-GFP remained within VLCs under both dark and BL BFA treatments, suggesting that phot1 and NPH3 play different roles in PIN2 localization. In conclusion, BL-induced root phototropism is based on the phot1/NPH3 signaling pathway, which stimulates the shootward auxin flux by modifying the subcellular targeting of PIN2 in the root apex transition zone.

Tissue-Autonomous Promotion of Palisade Cell Development by Phototropin 2 inArabidopsis

Light is an important environmental information source that plants use to modify their growth and development. Palisade parenchyma cells in leaves develop cylindrical shapes in response to blue light; however, the photosensory mechanism for this response has not been elucidated. In this study, we analyzed the palisade cell response in phototropin-deficient mutants. First, we found that two different light-sensing mechanisms contributed to the response in different proportions depending on the light intensity. One response observed under lower intensities of blue light was mediated exclusively by a blue light photoreceptor, phototropin 2 (PHOT2). Another response was elicited under higher intensities of light in a phototropin-independent manner. To determine the tissue in which PHOT2 perceives the light stimulus to regulate the response, green fluorescent protein (GFP)-tagged PHOT2 (P2G) was expressed under the control of tissue-specific promoters in the phot1 phot2 mutant background. The results revealed that the expression of P2G in the mesophyll, but not in the epidermis, promoted palisade cell development. Furthermore, a constitutively active C-terminal kinase fragment of PHOT2 fused to GFP (P2CG) promoted the development of cylindrical palisade cells in the proper direction without the directional cue provided by light. Hence, in response to blue light, PHOT2 promotes the development of cylindrical palisade cells along a predetermined axis in a tissue-autonomous manner.

Red Light, Phot1 and JAC1 Modulate Phot2-Dependent Reorganization of Chloroplast Actin Filaments and Chloroplast Avoidance Movement

The phototropin (phot)-dependent intracellular relocation of chloroplasts is a ubiquitous phenomenon in plants. We have previously revealed the involvement of a short cp-actin (chloroplast actin) filament-based mechanism in this movement. Here, the reorganization of cp-actin filaments during the avoidance movement of chloroplasts was analyzed in higher time resolution under blue GFP (green fluorescent protein) excitation light in an actin filament-visualized line of Arabidopsis thaliana. Under standard background red light of 89 μmol m(-2) s(-1), cp-actin filaments transiently disappeared at approximately 30 s and reappeared in a biased configuration on chloroplasts approximately 70 s after blue excitation light irradiation. The timing of biased cp-actin reappearance was delayed under the background of strong red light or in the absence of red light. Consistently, chloroplast movement was delayed under these conditions. In phot1 mutants, acceleration of both the disappearance and reappearance of cp-actin filaments occurred, indicating an inhibitory action of phot1 on reorganization of cp-actin filaments. Avoidance movements began sooner in phot1 than in wild-type plants. No reorganization of cp-actin filaments was seen in phot2 or phot1phot2 mutants lacking phot2, which is responsible for avoidance movements. Surprisingly, jac1 (j-domain protein required for chloroplast accumulation response 1) mutants, lacking the accumulation response, showed no avoidance movements under the whole-cell irradiation condition for GFP observation. Cp-actin filaments in jac1 did not show a biased distribution, with a small or almost no transient decrease in the number. These results indicate a close association between the biased distribution of cp-actin filaments and chloroplast movement. Further, JAC1 is suggested to function in the biased cp-actin filament distribution by regulating their appearance and disappearance.

Polarized by the Light

Light-dependent phosphorylation of the auxin efflux carrier PIN3 determines its trafficking pattern in the hypocotyl endodermis.

Evidence of physiological phototropin1 (phot1) action in response to UV-C illumination

Stem growth kinetics were measured in cucumber (Cucumis sativus) and Arabidopsis thaliana using highly-sensitive monitoring with 5-minute resolution, in darkness and in response to a short, single pulse of UV-C illumination. The results show that UV-C, like blue light, induces a rapid decrease in seedling growth rate. The fluence-response kinetics and time course were similar to the phototropin1 mediated response observed following a blue pulse. Arabidopsis seedlings were used to assess the genetic mechanism of this response. The phot1 mutant exhibited defects in stem growth rate inhibition, with sustained growth inhibition completely absent following specific treatments. The cryptochrome and phytochrome mutants exhibited responses comparable to wild type, suggesting that these receptor classes do not contribute to this response. The work demonstrates in two species that UV-C has an effect on a rapid plant photomorphogenic response and that the response is partially mediated by the phot1 photoreceptor.

Integration of phot1, phot2, and PhyB signalling in light-induced chloroplast movements

In Arabidopsis thaliana, chloroplasts move towards the periclinal cell walls upon exposure to low blue light intensities and to anticlinal walls under high light. The regulation of these chloroplast movements involves members of both the phototropin and phytochrome families of photoreceptors. Examination of fluence-rate response dependencies in phot1 and phot2 mutants revealed that although both photoreceptors are capable of inducing chloroplast accumulation under low-light conditions, the signals from these photoreceptors appear to be antagonistic. Chloroplast movements in wild-type plants were intermediate between those of the single phot mutants, consistent with each operating through separate signalling cascades. Mutants in phot2 showed transient chloroplast avoidance responses upon exposure to intense blue light, and slow but sustained chloroplast avoidance under intense white light, indicating that in the absence of phot2, phot1 is capable of generating both a low and a high-light response signal. Mutations in phytochrome B (phyB) caused an enhanced avoidance response at intermediate and high light intensities. Examination of phyB, phot1phyB, and phot2phyB mutants indicated that this enhancement is caused by PhyB inhibition of the high-light avoidance response in wild-type plants. In addition, our results suggest that the inhibition by PhyB is not exclusive to either of the phot1 or phot2 signalling pathways.

Actin Reorganization Underlies Phototropin-Dependent Positioning of Nuclei in Arabidopsis Leaf Cells

In epidermal and mesophyll cells of Arabidopsis (Arabidopsis thaliana) leaves, nuclei become relocated in response to strong blue light. We previously reported that nuclear positions both in darkness and in strong blue light are regulated by the blue light receptor phototropin2 in mesophyll cells. Here, we investigate the involvement of phototropin and the actin cytoskeleton in nuclear positioning in epidermal cells. Analysis of geometrical parameters revealed that, in darkness, nuclei were distributed near the center of the cell, adjacent to the inner periclinal wall, independent of cell shape. Dividing the anticlinal wall into concave, convex, and intermediate regions indicated that, in strong blue light, nuclei became relocated preferably to a concave region of the anticlinal wall, nearest the center of the cell. Mutant analyses verified that light-dependent nuclear positioning was regulated by phototropin2, while dark positioning of nuclei was independent of phototropin. Nuclear movement was inhibited by an actin-depolymerizing reagent, latrunculin B, but not by a microtubule-disrupting reagent, propyzamide. Imaging actin organization by immunofluorescence microscopy revealed that thick actin bundles, periclinally arranged parallel to the longest axis of the epidermal cell, were associated with the nucleus in darkness, whereas under strong blue light, the actin bundles, especially in the vicinity of the nucleus, became arranged close to the anticlinal walls. Light-dependent changes in the actin organization were clear in phot1 mutant but not in phot2 and phot1phot2 mutants. We propose that, in Arabidopsis, blue-light-dependent nuclear positioning is regulated by phototropin2-dependent reorganization of the actin cytoskeleton.

Blue Light-Induced Phototropism of Inflorescence Stems and Petioles is Mediated by Phototropin Family Members phot1 and phot2

Phototropin family photoreceptors, phot1 and phot2, in Arabidopsis thaliana control the blue light (BL)-mediated phototropic responses of the hypocotyl, chloroplast relocation movement and stomatal opening. Phototropic responses in dark-grown tissues have been well studied but those in de-etiolated green plants are not well understood. Here, we analyzed phototropic responses of inflorescence stems and petioles of wild-type and phototropin mutant plants of A. thaliana. Similar to the results obtained from dark-grown seedlings, inflorescence stems and petioles in wild-type and phot2 mutant plants showed phototropic bending towards low fluence BL, while in phot1 mutant plants, a high fluence rate of BL was required. phot1 phot2 double mutant plants did not show any phototropic responses even under very high fluence rates of BL. We further studied the photoreceptive sites for phototropic responses of stems and petioles by partial tissue irradiation. The whole part of the inflorescence stem is sensitive to BL and shows phototropism, but in the petiole only the irradiated abaxial side is sensitive. Similar to dark-grown etiolated seedlings, phot1 plays a major role in phototropic responses under weak light, but phot2 functions under high fluence rate conditions in green plants.

Blue Light‐induced Chloroplast Avoidance and Phototropic Responses Exhibit Distinct Dose Dependency of PHOTOTROPIN2 in Arabidopsis thaliana

PHOTOTROPIN2 (PHOT2) is a unique photoreceptor involved in chloroplast avoidance movement and also regulates blue light (BL) responses, such as phototropism and leaf flattening, together with PHOTOTROPIN1 (PHOT1) in Arabidopsis thaliana. Previous work showed that the defect of the phot2-1 mutant in chloroplast avoidance movement was a semidominant trait. In the present study, we examined PHOT2 dose dependency of BL responses using the phot1-5 phot2-1 double mutant expressing an AtPHOT2-GFP (P2G) fusion protein. Chloroplast avoidance and phototropic responses of P2G transgenic lines were enhanced in a manner dependent on the P2G levels, whereas the leaf flattening phenotype was simply complemented by P2G equivalent to the wild type (WT) PHOT2 level. The chloroplast avoidance velocity of P2G transgenic lines exhibited enhanced sensitivity to BL in comparison with WT. In contrast, the defect of the phototropic response was rescued by P2G expression equivalent only to the response of the phot1 mutant. These results collectively indicate that each BL response has distinct threshold levels of PHOT2 requirement.

Rice phot1a mutation reduces plant growth by affecting photosynthetic responses to light during early seedling growth

The aim of this work was to characterize the phot1 mutant of rice during early seedling growth in various light conditions. We isolated the rice T-DNA insertion mutant phot1a-1 and compared it to the Tos17 insertion mutant phot1a-2. When phot1a mutants were grown under WL (100) and BL (40 miccromol m(-2) s(-1)), they demonstrated a considerable reduction in photosynthetic capacity, which included decreased leaf CO(2) uptake and plant growth. Pigment analysis showed no significant difference between wild-type and mutants in the Chl a:b ratios, whereas in the latter, total concentration was reduced (a 2-fold decrease). Carotenoid contents of the mutants were also decreased considerably, implying the involvement of phot1a in pigment degradation. Deletion of phot1a showed higher contents of H(2)O(2) in leaves. Chloroplastic APX and SOD activities were lower in the mutants whereas the activities of cytosolic enzymes were increased. Immunoblotting indicated reduced accumulation of photosystem proteins (D1, D2, CP43, Lhca2, and PsaC) relative to the other light-harvesting complexes in the mutant. We conclude that the defect of Os Phot1a affects degradation of chlorophylls and carotenoids, and under photosynthetically active photon fluxes, mutation of phot1a results in loss of photosynthetic capacity owing to the damage of photosystems caused by elevated H(2)O(2) accumulation, leading to a reduction in plant growth.

Molecular basis of the functional specificities of phototropin 1 and 2

A blue-light photoreceptor in plants, phototropin, mediates phototropism, chloroplast relocation, stomatal opening, and leaf-flattening responses. Phototropin is divided into two functional moieties, the N-terminal photosensory and the C-terminal signaling moieties. Phototropin perceives light stimuli by the light, oxygen or voltage (LOV) domain in the N-terminus; the signal is then transduced intramolecularly to the C-terminal kinase domain. Two phototropins, phot1 and phot2, which have overlapping and distinct functions, exist in Arabidopsis thaliana. Phot1 mediates responses with higher sensitivity than phot2. Phot2 mediates specific responses, such as the chloroplast avoidance response and chloroplast dark positioning. To elucidate the molecular basis for the functional specificities of phot1 and phot2, we exchanged the N- and C-terminal moieties of phot1 and phot2, fused them to GFP and expressed them under the PHOT2 promoter in the phot1 phot2 mutant background. With respect to phototropism and other responses, the chimeric phototropin consisting of phot1 N-terminal and phot2 C-terminal moieties (P1n/2cG) was almost as sensitive as phot1; whereas the reverse combination (P2n/1cG) functioned with lower sensitivity. Hence, the N-terminal moiety mainly determined the sensitivity of the phototropins. Unexpectedly, both P1n/2cG and P2n/1cG mediated the chloroplast avoidance response, which is specific to phot2. Hence, chloroplast avoidance activity appeared to be suppressed specifically in the combination of N- and C-terminal moieties of phot1. Unlike the chloroplast avoidance response, chloroplast dark positioning was observed for P2G and P2n/1cG but not for P1G or P1n/2cG, suggesting that a specific structure in the N-terminal moiety of phot2 is required for this activity.

The chloroplast avoidance response decreases internal conductance to CO2 diffusion in Arabidopsis thaliana leaves

The relationship between chloroplast arrangement and diffusion of CO(2) from substomatal cavities to the chloroplast stroma was investigated in Arabidopsis thaliana. Chloroplast position was manipulated by varying the amount of blue light and by cytochalasin D (CytD) treatment. We also investigated two chloroplast positioning mutants. Chloroplast arrangement was assessed by the surface area of chloroplasts adjacent to intercellular airspaces (S(c)). Although it has been previously shown that long-term acclimation to high light is linked with a large S(c), we found that the short-term chloroplast avoidance response reduces S(c). This effect was not apparent in the blue-light-insensitive phot2 mutant, which did not show the avoidance response. As expected, the smaller S(c) induced by the avoidance response was coupled to a similar decrease in internal conductance. This reduction in internal conductance resulted in an increased limitation of the rate of photosynthesis. The limiting effect of S(c) on internal conductance and photosynthesis was also shown in chup1, a mutant with a constant small S(c) as the result of an unusual chloroplast arrangement. We conclude that chloroplast movements in A. thaliana can rapidly alter leaf morphological parameters, and this has significant consequences for the diffusion of CO(2) through the mesophyll.

Mapping of the phosphorylation sites on the phototropic signal transducer, NPH3

The phototropic response in Arabidopsis thaliana is initiated by the blue-light photoreceptors, phototropin (phot)1 and phot2. A recent study has shown that one of the phototropic signal transducers, NPH3, is phosphorylated under dark conditions and dephosphorylated by blue-light irradiation. To further understand the function of phosphorylation and dephosphorylation of NPH3 during this phototropic response, we have mapped the phosphorylation sites of NPH3 in our current study. The NPH3 protein has at least three phosphorylation sites at serine residues, Ser212, Ser222, and Ser236, and these sites are dephosphorylated by blue-light irradiation. By immunoblotting analysis, these phosphorylation sites in phot1 mutants are not dephosphorylated following blue-light irradiation at both low and high fluence rates, even though such irradiations induce the phot2-dependent phototropic response in phot1. These results suggest that the dephosphorylated NPH3 is involved in the phot1-dependent phototropic response and is not essential for the phot2-dependent phototropic response. We generated two types of transgenic nph3 plants expressing a NPH3 S212A/S222A/S232A/S236A protein and a NPH3 Δ212–238 protein in which the phosphorylation region is deleted, and assessed the phototropic phenotype of these. Based upon our present findings, we discuss the role of dephosphorylated and phosphorylated NPH3 in mediating the phototropic response.

An Inositol Polyphosphate 5-Phosphatase Functions in PHOTOTROPIN1 Signaling in Arabidopis by Altering Cytosolic Ca2+

Inositol polyphosphate 5-phosphatase (5PTase) is a key enzyme in the phosphatidylinositol metabolic pathway, which plays critical roles in a number of cellular processes in plants. Our previous work implicated the role of 5PTase13, which encodes a WD40-containing type II 5PTase, in hormone-mediated cotyledon vein development. Here, we show that 5PTase13 is also involved in blue light responses in Arabidopsis thaliana. Compared with that in darkness, the expression of 5PTase13 was suppressed by blue light irradiation, and disruption of the gene resulted in shortened hypocotyls and expanded cotyledons. Genetic analysis showed that 5PTase13 acted independently from CRYPTOCHROME1 and CONSTITUTIVE PHOTOMORPHOGENIC1 but interacted functionally with PHOTOTROPIN1 (PHOT1). The expression level of 5PTase13 was significantly enhanced in phot1 single or phot1 phot2 double mutants under blue light, and suppression of 5PTase13 expression rescued the elongated hypocotyls in the phot1 or phot1 phot2 mutants. Further analysis showed that the blue light-induced elevation of cytosolic Ca2+ was inhibited in the phot1 mutant but enhanced in the 5pt13 mutant, suggesting that 5PTase13 antagonizes PHOT1-mediated effects on calcium signaling under blue light.

Multiple interactions between cryptochrome and phototropin blue-light signalling pathways in Arabidopsis thaliana

Higher plants contain two structurally unrelated flavoprotein blue-light photoreceptors, the cryptochromes and the phototropins, which mediate largely distinct response pathways. Cryptochromes regulate plant development and photomorphogenesis whereas phototropins are primarily implicated in photomovement responses such as phototropism and chloroplast relocation. In the present study we identify interactions between cryptochromes and phototropins in several photoresponses of Arabidopsis thaliana. Cryptochromes are shown to exert a positive effect on phototropic curvature under long-term irradiation conditions. Specifically, in a phot1-deficient genetic background (phot1 mutant), curvature is reduced in the absence of cryptochromes, particularly at wavelengths where cryptochromes show preferential absorption. Phototropins in turn exert a small promotive effect on such cryptochrome-mediated responses as hypocotyl elongation and anthocyanin accumulation. These effects are apparent in a cryptochrome-deficient (cry1cry2 mutant) genetic background. In addition to positive interactions between signalling pathways, we demonstrate that the cryptochromes also exert a negative regulatory effect. Levels of phot1 protein decrease in blue light as a function of cryptochrome photoreceptor activation. This negative regulation occurs in part at the level of phot1 transcription but may also involve post-transcriptional mechanisms. These two classes of photoreceptor thereby reciprocally modulate their overall responsivity to blue light through multiple forms of interaction.

Leaf Positioning of Arabidopsis in Response to Blue Light

Appropriate leaf positioning is essential for optimizing photosynthesis and plant growth. However, it has not been elucidated how green leaves reach and maintain their position for capturing light. We show here the regulation of leaf positioning under blue light stimuli. When 1-week-old Arabidopsis seedlings grown under white light were transferred to red light (25 micromol m(-2) s(-1)) for 5 d, new petioles that appeared were almost horizontal and their leaves were curled and slanted downward. However, when a weak blue light from above (0.1 micromol m(-2) s(-1)) was superimposed on red light, the new petioles grew obliquely upward and the leaves were flat and horizontal. The leaf positioning required both phototropin1 (phot1) and nonphototropic hypocotyl 3 (NPH3), and resulted in enhanced plant growth. In an nph3 mutant, neither optimal leaf positioning nor leaf flattening by blue light was found, and blue light-induced growth enhancement was drastically reduced. When blue light was increased from 0.1 to 5 micromol m(-2) s(-1), normal leaf positioning and leaf flattening were induced in both phot1 and nph3 mutants, suggesting that phot2 signaling became functional and that the signaling was independent of phot1 and NPH3 in these responses. When plants were irradiated with blue light (0.1 micromol m(-2) s(-1)) from the side and red light from above, the new leaves became oriented toward the source of blue light. When we transferred these plants to both blue light and red light from above, the leaf surface changed its orientation to the new blue light source within a few hours, whereas the petioles initially were unchanged but then gradually rotated, suggesting the plasticity of leaf positioning in response to blue light. We showed the tissue expression of NPH3 and its plasma membrane localization via the coiled-coil domain and the C-terminal region. We conclude that NPH3-mediated phototropin signaling optimizes the efficiency of light perception by inducing both optimal leaf positioning and leaf flattening, and enhances plant growth.

Phototropin Mediated Relocation of Myosins in Arabidopsis thaliana

Background information. The mechanism of the light-dependent movements of chloroplasts is based on actin and myosin but its details are largely unknown. The movements are activated by blue light in terrestrial angiosperms. The aim of the present study was to determine the role of myosin associated with the chloroplast surface in the light-induced chloroplast responses in Arabidopsis thaliana. The localization of myosins was investigated under blue light intensities generating avoidance and accumulation responses of chloroplasts. The localization was compared in wild type plants and in phot2 mutant lacking the avoidance response.Results. Wild type and phot2 mutant plants were irradiated with strong (36 μEm-2s-1) and/or weak (0,8 μEm-2s-1) blue light. The leaf tissue was immunolabeled with anti-myosin antibodies. Different arrangements of myosins were observed in the mesophyll depending on the fluence rate in wild type plants. In tissue irradiated with weak blue light myosins were associated with chloroplast envelopes. In contrast, in tissue irradiated with strong blue light chloroplasts were almost myosin-free. The effect did not occur in red light and in the phot2 mutant.Conclusions. Myosin displacement is blue light specific, i.e. it is associated with the activation of a specific blue-light photoreceptor. We suggest that the reorganization of myosins is essential for chloroplast movement. Myosins appear to be the final step of the signal transduction pathway starting with phototropin2 and leading to chloroplast movements.