UV RESISTANCE LOCUS 8 Signaling Research Articles

Nitric oxide is a key part of the UV-B-induced photomorphogenesis in Arabidopsis

Ultraviolet- B (UV-B) radiation exerts a dual effect on plants. While high doses induce stress, lower doses act as a signal. The sensing of UV-B triggers a photomorphogenic response, causing changes like the hypocotyl shortening. This response is strongly regulated by the UV-B photoreceptor UV RESISTANCE LOCUS 8 (UVR8). Although nitric oxide (NO) was proven to inhibit hypocotyl elongation in Arabidopsis thaliana, its role in UV-B-induced photomorphogenesis remains unclear. By using the NO probe DAF-FM DA, we found that UV-B radiation induced the accumulation of NO in the hypocotyl. This event is dependent on the presence of UVR8. Moreover, the NO donor S-nitrosoglutathione (GSNO), restored the WT phenotype when the Arabidopsis line lacking UVR8 was grown in the presence of UV-B. Also, we found that DELLA proteins were necessary for GSNO sensing in UV-B-induced photomorphogenesis. We showed that, through DELLAs, GSNO negatively controlled the levels of gibberellin (GA) bioprocessing-related genes. Therefore, a downregulation of GA signaling by NO is proposed. On the other hand, GSNO supplementation before UV-B treatment inhibited the induction of genes of the UVR8 signaling pathway: HY5, CHS, and PHR1. This resulted in augmented DNA damage in short exposure to UV-B scenarios. Altogether, these results point to a complex regulation by NO of the UV-B-induced photomorphogenesis.

Arabidopsis B-box transcription factors BBX20-22 promote UVR8 photoreceptor-mediated UV-B responses.

SUMMARYPlants undergo photomorphogenic development in the presence of light. Photomorphogenesis is repressed by the E3 ubiquitin ligase CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1), which binds to substrates through their valine–proline (VP) motifs. The UV RESISTANCE LOCUS 8 (UVR8) photoreceptor senses UV‐B and inhibits COP1 through the cooperative binding of its own VP motif and photosensing core to COP1, thereby preventing COP1 binding to substrates, including the basic leucine zipper (bZIP) transcriptional regulator ELONGATED HYPOCOTYL 5 (HY5). As a key promoter of visible light and UV‐B photomorphogenesis, HY5 requires coregulators for its function. The B‐box family transcription factors BBX20–BBX22 were recently described as HY5 rate‐limiting coactivators under red light, but their role in UVR8 signaling was unknown. Here we describe a hypermorphic bbx21‐3D mutant with enhanced photomorphogenesis, carrying a proline‐to‐leucine mutation at position 314 in the VP motif that impairs the interaction with and regulation by COP1. We show that BBX21 and BBX22 are UVR8‐dependently stabilized after UV‐B exposure, which is counteracted by a repressor induced by HY5/BBX activity. bbx20 bbx21 bbx22 mutants under UV‐B are impaired in hypocotyl growth inhibition, photoprotective pigment accumulation and the expression of several HY5‐dependent genes under continuous UV‐B, but the immediate induction of marker genes after exposure to UV‐B remains surprisingly rather unaffected. We conclude that BBX20–BBX22 contribute to HY5 activity in a subset of UV‐B responses, but that additional, presently unknown, coactivators for HY5 are functional in early UVR8 signaling.

Transcriptome and metabolome analyses revealed that narrowband 280 and 310\xa0nm UV-B induce distinctive responses in Arabidopsis

In plants, the UV-B photoreceptor UV RESISTANCE LOCUS8 (UVR8) perceives UV-B and induces UV-B responses. UVR8 absorbs a range of UV-B (260–335 nm). However, the responsiveness of plants to each UV-B wavelength has not been intensively studied so far. Here, we performed transcriptome and metabolome analyses of Arabidopsis using UV light emitting diodes (LEDs) with peak wavelengths of 280 and 310 nm to investigate the differences in the wavelength-specific UV-B responses. Irradiation with both UV-LEDs induced gene expression of the transcription factor ELONGATED HYPOCOTYL 5 (HY5), which has a central role in the UVR8 signaling pathway. However, the overall transcriptomic and metabolic responses to 280 and 310 nm UV-LED irradiation were different. Most of the known UV-B-responsive genes, such as defense-related genes, responded only to 280 nm UV-LED irradiation. Lipids, polyamines and organic acids were the metabolites most affected by 280 nm UV-LED irradiation, whereas the effect of 310 nm UV-LED irradiation on the metabolome was considerably less. Enzymatic genes involved in the phenylpropanoid pathway upstream in anthocyanin biosynthesis were up-regulated only by 280 nm UV-LED irradiation. These results revealed that the responsivenesses of Arabidopsis to 280 and 310 nm UV-B were significantly different, suggesting that UV-B signaling is mediated by more complex pathways than the current model.

UVR8-mediated inhibition of shade avoidance involves HFR1 stabilization in Arabidopsis.

Sun-loving plants perceive the proximity of potential light-competing neighboring plants as a reduction in the red:far-red ratio (R:FR), which elicits a suite of responses called the "shade avoidance syndrome" (SAS). Changes in R:FR are primarily perceived by phytochrome B (phyB), whereas UV-B perceived by UV RESISTANCE LOCUS 8 (UVR8) elicits opposing responses to provide a counterbalance to SAS, including reduced shade-induced hypocotyl and petiole elongation. Here we show at the genome-wide level that UVR8 broadly suppresses shade-induced gene expression. A subset of this gene regulation is dependent on the UVR8-stabilized atypical bHLH transcription regulator LONG HYPOCOTYL IN FAR-RED 1 (HFR1), which functions in part redundantly with PHYTOCHROME INTERACTING FACTOR 3-LIKE 1 (PIL1). In parallel, UVR8 signaling decreases protein levels of the key positive regulators of SAS, namely the bHLH transcription factors PHYTOCHROME INTERACTING FACTOR 4 (PIF4) and PIF5, in a COP1-dependent but HFR1-independent manner. We propose that UV-B antagonizes SAS via two mechanisms: degradation of PIF4 and PIF5, and HFR1- and PIL1-mediated inhibition of PIF4 and PIF5 function. This work highlights the importance of typical and atypical bHLH transcription regulators for the integration of light signals from different photoreceptors and provides further mechanistic insight into the crosstalk of UVR8 signaling and SAS.

Degradation of the transcription factors PIF4 and PIF5 under UV-B promotes UVR8-mediated inhibition of hypocotyl growth in Arabidopsis.

SummaryInhibition of hypocotyl growth is a well‐established UV‐B‐induced photomorphogenic response that is mediated by the UV‐B photoreceptor UV RESISTANCE LOCUS 8 (UVR8). However, the molecular mechanism by which UVR8 signaling triggers inhibition of hypocotyl growth is poorly understood. The bZIP protein ELONGATED HYPOCOTYL 5 (HY5) functions as the main positive regulatory transcription factor in the UVR8 signaling pathway, with HY5‐HOMOLOG (HYH) playing a minor role. However, here we demonstrate that hy5 hyh double mutants maintain significant UVR8‐dependent hypocotyl growth inhibition. We identify UVR8‐dependent inhibition of the activities of bHLH transcription factors PHYTOCHROME INTERACTING FACTOR 4 (PIF4) and PIF5 as part of the UVR8 signaling pathway, which results in inhibition of hypocotyl growth. The UVR8‐mediated repression of several hypocotyl elongation‐related genes is independent of HY5 and HYH but largely associated with UVR8‐dependent degradation of PIF4 and PIF5, a process that consequently diminishes PIF4/5 target promoter occupancy. Taken together, our data indicate that UVR8‐mediated inhibition of hypocotyl growth involves degradation of PIF4 and PIF5. These findings contribute to our mechanistic understanding of UVR8‐induced photomorphogenesis and further support the function of PIFs as integrators of different photoreceptor signaling pathways.

Regulation of Arabidopsis gene expression by low fluence rate UV-B independently of UVR8 and stress signaling

UV-B exposure of plants regulates expression of numerous genes concerned with various responses. Sudden exposure of non-acclimated plants to high fluence rate, short wavelength UV-B induces expression via stress-related signaling pathways that are not specific to the UV-B stimulus, whereas low fluence rates of UV-B can regulate expression via the UV-B photoreceptor UV RESISTANCE LOCUS 8 (UVR8). However, there is little information about whether non-stressful, low fluence rate UV-B treatments can activate gene expression independently of UVR8. Here, transcriptomic analysis of wild-type and uvr8 mutant Arabidopsis exposed to low fluence rate UV-B showed that numerous genes were regulated independently of UVR8. Moreover, nearly all of these genes were distinct to those induced by stress treatments. A small number of genes were expressed at all UV-B fluence rates employed and may be concerned with activation of eustress responses that facilitate acclimation to changing conditions. Expression of the gene encoding the transcription factor ARABIDOPSIS NAC DOMAIN CONTAINING PROTEIN 13 (ANAC13) was studied to characterise a low fluence rate, UVR8-independent response. ANAC13 is induced by as little as 0.1 μmol m−2 s−1 UV-B and its regulation is independent of components of the canonical UVR8 signaling pathway COP1 and HY5/HYH. Furthermore, UV-B induced expression of ANAC13 is independent of the photoreceptors CRY1, CRY2, PHOT1 and PHOT2 and phytochromes A, B, D and E. ANAC13 expression is induced over a range of UV-B wavelengths at low doses, with maximum response at 310 nm. This study provides a basis for further investigation of UVR8 and stress independent, low fluence rate UV-B signaling pathway(s).

Origin and Evolution of Core Components Responsible for Monitoring Light Environment Changes during Plant Terrestrialization

Light serves as the source of energy as well as an information signal for photosynthetic plants. During evolution, plants have acquired the ability to monitor environmental light radiation and adjust their developmental patterns to optimally utilize light energy for photosynthesis. The mechanisms of light perception and signal transduction have been comprehensively studied in past decades, mostly in a few model plants, including Arabidopsis thaliana. However, systematic analyses of the origin and evolution of core components involved in light perception and signaling are still lacking. In this study, we took advantage of the recently sequenced genomes and transcriptomes covering all the main Archaeplastida clades in the public domain to identify orthologous genes of core components involved in light perception and signaling and to reconstruct their evolutionary history. Our analyses suggested that acclimation to different distribution of light quality in new environments led to the origination of specific light signaling pathways in plants. The UVR8 (UV Resistance Locus 8) signaling pathway originated during the movement of plants from the deeper sea to shallow water and enabled plants to deal with ultraviolet B light (UV-B). After acquisition of UV-B adaptation, origination of the phytochrome signaling pathway helped plants to colonize water surface where red light became the prominent light energy source. The seedling emergence pathway, which is mediated by a combination of light and phytohormone signals that orchestrate plant growth pattern transitions, originated before the emergence of seed plants. Although cryptochromes and some key components of E3 ubiquitin ligase systems already existed before the divergence of the plant and animal kingdoms, the coevolution and optimization of light perception and downstream signal transduction components, including key transcription factors and E3 ubiquitin ligase systems, are evident during plant terrestrialization.

REPRESSOR OF ULTRAVIOLET-B PHOTOMORPHOGENESIS function allows efficient phototropin mediated ultraviolet-B phototropism in etiolated seedlings

Ultraviolet B (UV-B) light is a part of the solar radiation which has significant effects on plant morphology, even at low doses. In Arabidopsis, many of these morphological changes have been attributed to a specific UV-B receptor, UV resistance locus 8 (UVR8). Recent findings showed that next to phototropin regulated phototropism, UVR8 mediated signaling is able of inducing directional bending towards UV-B light in etiolated seedlings of Arabidopsis, in a phototropin independent manner. In this study, kinetic analysis of phototropic bending was used to evaluate the relative contribution of each of these pathways in UV-B mediated phototropism. Diminishing UV-B light intensity favors the importance of phototropins. Molecular and genetic analyses suggest that UV-B is capable of inducing phototropin signaling relying on phototropin kinase activity and regulation of NPH3. Moreover, enhanced UVR8 responses in the UV-B hypersensitive rup1rup2 mutants interferes with the fast phototropin mediated phototropism. Together the data suggest that phototropins are the most important receptors for UV-B induced phototropism in etiolated seedlings, and a RUP mediated negative feedback pathway prevents UVR8 signaling to interfere with the phototropin dependent response.

COP1 is required for UV-B–induced nuclear accumulation of the UVR8 photoreceptor

The UV-B photoreceptor UV RESISTANCE LOCUS 8 (UVR8) promotes UV-B acclimation and tolerance in Arabidopsis thaliana UVR8 localizes to both cytosol and nucleus, but its main activity is assumed to be nuclear. UV-B photoreception stimulates nuclear accumulation of UVR8 in a presently unknown manner. Here, we show that CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) is required for UV-B-induced nuclear accumulation of UVR8, but bypassing the COP1 requirement for UVR8 nuclear accumulation did not rescue the cop1 mutant UV-B phenotype. Using a glucocorticoid receptor (GR)-based fusion protein system to conditionally localize GR-UVR8 to the nucleus, we have demonstrated that both photoactivation and nuclear localization of UVR8 are required for UV-B-induced photomorphogenic responses. In contrast, there was no UV-B response when UV-B-activated UVR8 was artificially retained in the cytosol. In agreement with a predominantly nuclear activity, constitutively active UVR8(W285A) accumulated in the nucleus also in the absence of UV-B. Furthermore, GR-COP1 expression lines suggested that UV-B-activated UVR8 can be coimported into the nucleus by COP1. Our data strongly support localization of UVR8 signaling in the nucleus and a dual role for COP1 in the regulation of UV-B-induced UVR8 nuclear accumulation and in UVR8-mediated UV-B signaling.

Ultraviolet-B radiation stimulates downward leaf curling in Arabidopsis thaliana

Plants are very well adapted to growth in ultraviolet-B (UV-B) containing light. In Arabidopsis thaliana, many of these adaptations are mediated by the UV-B receptor UV RESISTANCE LOCUS 8 (UVR8). Using small amounts of supplementary UV-B light, we observed changes in the shape of rosette leaf blades. Wild type plants show more pronounced epinasty of the blade edges, while this is not the case in uvr8 mutant plants. The UVR8 effect thus mimics the effect of phytochrome (phy) B in red light. In addition, a meta-analysis of transcriptome data indicates that the UVR8 and phyB signaling pathways have over 70% of gene regulation in common. Moreover, in low levels of supplementary UV-B light, mutant analysis revealed that phyB signaling is necessary for epinasty of the blade edges. Analysis of auxin levels and the auxin signal reporter DR5::GUS suggest that the epinasty relies on altered auxin distribution, keeping auxin at the leaf blade edges in the presence of UV-B. Together, our results suggest a co-action of phyB and UVR8 signaling, with auxin as a downstream factor.

Ultraviolet-B-induced stomatal closure in Arabidopsis is regulated by the UV RESISTANCE LOCUS8 photoreceptor in a nitric oxide-dependent mechanism.

UV RESISTANCE LOCUS8 (UVR8) signaling involves CONSTITUTIVELY PHOTOMORPHOGENIC1, the ELONGATED HYPOCOTYL5 (HY5) transcription factor, and the closely related HY5 HOMOLOG. Some UV-B responses mediated by UVR8 are also regulated by nitric oxide (NO), a bioactive molecule that orchestrates a wide range of processes in plants. In this study, we investigated the participation of the UVR8 pathway and its interaction with NO in UV-B-induced stomatal movements in Arabidopsis (Arabidopsis thaliana). Stomata in abaxial epidermal strips of Arabidopsis ecotype Landsberg erecta closed in response to increasing UV-B fluence rates, with maximal closure after 3-h exposure to 5.46 μmol m⁻² s⁻¹ UV-B. Both hydrogen peroxide (H₂O₂) and NO increased in response to UV-B, and stomatal closure was maintained by NO up to 24 h after the beginning of exposure. Stomata of plants expressing bacterial NO dioxygenase, which prevents NO accumulation, did not close in response to UV-B, although H₂O₂ still increased. When the uvr8-1 null mutant was exposed to UV-B, stomata remained open, irrespective of the fluence rate. Neither NO nor H₂O₂ increased in stomata of the uvr8-1 mutant. However, the NO donor S-nitrosoglutathione induced closure of uvr8-1 stomata to the same extent as in the wild type. Experiments with mutants in UVR8 signaling components implicated CONSTITUTIVELY PHOTOMORPHOGENIC1, HY5, and HY5 HOMOLOG in UV-B-induced stomatal closure. This research provides evidence that the UVR8 pathway regulates stomatal closure by a mechanism involving both H₂O₂ and NO generation in response to UV-B exposure.

UV‐B Action Spectrum for UVR8‐Mediated HY5 Transcript Accumulation in Arabidopsis

Arabidopsis thaliana UV RESISTANCE LOCUS8 (UVR8) is a UV-B-specific signaling component that mediates low fluence, photomorphogenic responses to UV-B. It is required for UV-B-induced expression of the gene encoding the ELONGATED HYPOCOTYL5 (HY5) transcription factor. HY5 is a key effector of responses mediated by UVR8. In mature leaf tissue, HY5 transcript accumulation occurred rapidly in response to a brief UV-B treatment and no induction was observed in a uvr8 mutant over a broad range of UV wavelengths. In response to monochromatic light, maximal transcript accumulation occurred in wild-type plants at wavelengths 280-300 nm. HY5 transcript accumulation showed reciprocity between the fluence rate and duration of UV-B exposure, and on this basis conditions were chosen to generate an action spectrum for the UVR8 signaling pathway. Dose-response curves were produced for a range of UV wavelengths using 20 min exposure to UV and harvesting tissue 2 h after the start of illumination. Experiments using mutants defective in sinapate ester and flavonoid biosynthesis indicated that the presence of UV-absorbing compounds did not affect the construction of an action spectrum under the conditions employed. The action spectrum for the induction of HY5 by the UVR8 pathway showed a main peak at 280 nm with a smaller peak at 300 nm. The data are discussed in relation to the proposed mechanisms of UV-B photoreception.