Nuclear accumulation of rice UV-B photoreceptors is UV-B- and OsCOP1-independent for UV-B responses.

In plants, the conserved plant-specific photoreceptor UV RESISTANCE LOCUS 8 (UVR8) perceives ultraviolet-B (UV-B) light and mediates UV-B-induced photomorphogenesis and stress acclimation. In this study, we reveal that UV-B light treatment shortens seedlings, increases stem thickness, and enhances UV-B stress tolerance in rice (Oryza sativa) via its two UV-B photoreceptors OsUVR8a and OsUVR8b. Although the rice and Arabidopsis (Arabidopsis thaliana) UVR8 (AtUVR8) photoreceptors all form monomers in response to UV-B light, OsUVR8a, and OsUVR8b function is only partially conserved with respect to AtUVR8 in UV-B-induced photomorphogenesis and stress acclimation. UV-B light and CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) promote the nuclear accumulation of AtUVR8; by contrast, OsUVR8a and OsUVR8b constitutively localize to the nucleus via their own nuclear localization signals, independently of UV-B light and the RING-finger mutation of OsCOP1. We show that OsCOP1 negatively regulates UV-B responses, and shows weak interaction with OsUVR8s, which is ascribed to the N terminus of OsCOP1, which is conserved in several monocots. Furthermore, transcriptome analysis demonstrates that UV-B-responsive gene expression differs globally between Arabidopsis and rice, illuminating the evolutionary divergence of UV-B light signaling pathways between monocot and dicot plants.

Photoperiod Genes Contribute to Daylength-Sensing and Breeding in Rice.

Rice (Oryza sativa L.), one of the most important food crops worldwide, is a facultative short-day (SD) plant in which flowering is modulated by seasonal and temperature cues. The photoperiodic molecular network is the core network for regulating flowering in rice, and is composed of photoreceptors, a circadian clock, a photoperiodic flowering core module, and florigen genes. The Hd1-DTH8-Ghd7-PRR37 module, a photoperiodic flowering core module, improves the latitude adaptation through mediating the multiple daylength-sensing processes in rice. However, how the other photoperiod-related genes regulate daylength-sensing and latitude adaptation remains largely unknown. Here, we determined that mutations in the photoreceptor and circadian clock genes can generate different daylength-sensing processes. Furthermore, we measured the yield-related traits in various mutants, including the main panicle length, grains per panicle, seed-setting rate, hundred-grain weight, and yield per panicle. Our results showed that the prr37, elf3-1 and ehd1 mutants can change the daylength-sensing processes and exhibit longer main panicle lengths and more grains per panicle. Hence, the PRR37, ELF3-1 and Ehd1 locus has excellent potential for latitude adaptation and production improvement in rice breeding. In summary, this study systematically explored how vital elements of the photoperiod network regulate daylength sensing and yield traits, providing critical information for their breeding applications.

Gradual daylength sensing coupled with optimum cropping modes enhances multi-latitude adaptation of rice and maize.

To expand crop planting areas, reestablishment of crop latitude adaptation based on genetic variation in photoperiodic genes can be performed, but it is quite time consuming. By contrast, a crop variety that already exhibits multi-latitude adaptation has the potential to increase its planting areas to be more widely and quickly available. However, the importance and potential of multi-latitude adaptation of crop varieties have not been systematically described. Here, combining daylength-sensing data with the cropping system of elite rice and maize varieties, we found that varieties with gradual daylength sensing coupled with optimum cropping modes have an enhanced capacity for multi-latitude adaptation in China. Furthermore, this multi-latitude adaptation expanded their planting areas and indirectly improved China's nationwide rice and maize unit yield. Thus, coupling the daylength-sensing process with optimum cropping modes to enhance latitude adaptability of excellent varieties represents an exciting approach for deploying crop varieties with the potential to expand their planting areas and quickly improve nationwide crop unit yield in developing countries.

Genome-wide prediction of activating regulatory elements in rice by combining STARR-seq with FACS.

Self-transcribing active regulatory region sequencing (STARR-seq) is widely used to identify enhancers at the whole-genome level. However, whether STARR-seq works as efficiently in plants as in animal systems remains unclear. Here, we determined that the traditional STARR-seq method can be directly applied to rice (Oryza sativa) protoplasts to identify enhancers, though with limited efficiency. Intriguingly, we identified not only enhancers but also constitutive promoters with this technique. To increase the performance of STARR-seq in plants, we optimized two procedures. We coupled fluorescence activating cell sorting (FACS) with STARR-seq to alleviate the effect of background noise, and we minimized PCR cycles and retained duplicates during prediction, which significantly increased the positive rate for activating regulatory elements (AREs). Using this method, we determined that AREs are associated with AT-rich regions and are enriched for a motif that the AP2/ERF family can recognize. Based on GC content preferences, AREs are clustered into two groups corresponding to promoters and enhancers. Either AT- or GC-rich regions within AREs could boost transcription. Additionally, disruption of AREs resulted in abnormal expression of both proximal and distal genes, which suggests that STARR-seq-revealed elements function as enhancers in vivo. In summary, our work provides a promising method to identify AREs in plants.

A Daylength Recognition Model of Photoperiodic Flowering.

The photoperiodic flowering pathway is crucial for plant development to synchronize internal signaling events and external seasons. One hundred years after photoperiodic flowering was discovered, the underlying core signaling network has been elucidated in model plants such as Arabidopsis (Arabidopsis thaliana), rice (Oryza sativa), and soybean (Glycine max). Here, we review the progress made in the photoperiodic flowering area and summarize previously accepted photoperiodic flowering models. We then introduce a new model based on daylength recognition by florigen. By determining the expression levels of the florigen gene, this model can assess the mechanism of daylength sensing and crop latitude adaptation. Future applications of this model under the constraints of global climate change are discussed.

Chromosomal Looping-Based Expression Activation System in Yeast.

There is growing evidence that 3D genome organization is a universal and significant mechanism of gene expression regulation. Tools to manipulate long-range DNA interactions can advance the accurate control of chromosomal architecture. However, simple eukaryotic systems available to engineer chromosomal looping and gene expression are very limited. This study has developed a tool designated as chromosomal looping-based expression activation system in yeast (CLEASY). Based on a modified yeast chromosome, it consists of conditionally interacting proteins, distal transcriptional regulatory elements, and a reporter gene. Exogenous chemical or light exposure induces the protein interaction, and results in the proximity of distal regulatory elements bound by these interacting proteins, and ultimately activates the reporter gene. In addition to this controllable induction, this system is compatible with the bivalent Cas9 complexes and their guide RNAs, to ensure target specificity and variability. Therefore, CLEASY can be utilized as a simplified eukaryotic model to engineer DNA looping machinery, and potentially serves as a fast platform to investigate looping mechanism and effective molecules.

Different response modes and cooperation modulations of blue-light receptors in photomorphogenesis.

Cryptochromes photoreceptors, CRY1 and CRY2 in Arabidopsis, mediate blue light responses in plants and metazoa. The signalling interactions underlying photomorphogenesis of cryptochromes action have been extensively studied in experiment, expecting a systematical analysis of the dynamic mechanisms of photosensory signalling network from a global view. In this study, we developed a signalling network model to quantitatively investigate the different response modes and cooperation modulations on photomorphogenesis for CRY1 and CRY2 under blue light. The model shows that the different modes of time-dependent and fluence-rate-dependent phosphorylations for CRY1 and CRY2 are originated from their different phosphorylation rates and degradation rates. Our study indicates that, due to the strong association between blue-light inhibitor of cryptochromes (BIC) and CRY2, BIC negatively modulates CRY2 phosphorylation, which was confirmed by our experiment. The experiment also validated the model prediction that the time-dependent BIC-CRY1 and the fluence-rate-dependent BIC-CRY2 are both bell-shaped under blue light. Importantly, the model proposes that the COP1-SPA abundance can strongly inhibit the phosphorylation response of CRY2, resulting in the positive regulation of CRY2 phosphorylation by CRY1 through COP1-SPA. The model also predicts that the CRY1-HY5 axis, rather than CRY2-HY5 pathway, plays a dominant role in blue-light-dependent photomorphogenesis.

Forecasting rice latitude adaptation through a daylength-sensing-based environment adaptation simulator.

Global climate change necessitates crop varieties with good environmental adaptability. As a proxy for climate adaptation, crop breeders could select for adaptability to different latitudes, but the lengthy procedures for that slow development. Here, we combined molecular technologies with a streamlined in-house screening method to facilitate rapid selection for latitude adaptation. We established the daylength-sensing-based environment adaptation simulator (DEAS) to assess rice latitude adaptation status via the transcriptional dynamics of florigen genes at different latitudes. The DEAS predicted the florigen expression profiles in rice varieties with high accuracy. Furthermore, the DEAS showed potential for application in different crops. Incorporating the DEAS into conventional breeding programmes would help to develop cultivars for climate adaptation.

Optogenetic tools controlled by ultraviolet-B light.

Decades of genetic, molecular and biochemical studies in plants have provided foundational knowledge about light sensory proteins and led to their application in synthetic biology. Optogenetic tools take advantage of the light switchable activity of plant photoreceptors to control intracellular signaling pathways. The recent discovery of the UV-B photoreceptor UV RESISTANCE LOCUS 8 in the model plant Arabidopsis thaliana opens up new avenues for light-controllable methodologies. In this review, we discuss current developments in optogenetic control by UV-B light and its signaling components, as well as rational considerations in the design and applications of UV-B-based optogenetic tools.

Coordinated Transcriptional Regulation by the UV-B Photoreceptor and Multiple Transcription Factors for Plant UV-B Responses.

Non-damaging ultraviolet B (UV-B) light promotes photomorphogenic development and stress acclimation through UV-B-specific signal transduction in Arabidopsis. UV-B irradiation induces monomerization and nuclear translocation of the UV-B photoreceptor UV RESISTANCE LOCUS 8 (UVR8). However, it is not clear how the nuclear localization of UVR8 leads to changes in global gene expression. Here, we reveal that nuclear UVR8 governs UV-B-responsive transcriptional networks in concert with several previously known transcription factors, including ELONGATED HYPOCOTYL 5 (HY5) and PHYTOCHROME INTERACTING FACTOR 4 (PIF4). Based on the transcriptomic analysis, we identify MYB13 as a novel positive regulator in UV-B-induced cotyledon expansion and stress acclimation. MYB13 is UV-B inducible and is predominantly expressed in the cotyledons. Our results demonstrate that MYB13 protein functions as a transcription factor to regulate the expression of genes involved in auxin response and flavonoid biosynthesis through direct binding with their promoters. In addition, photoactivated UVR8 interacts with MYB13 in a UV-B-dependent manner and differentially modulates the affinity of MYB13 with its targets. Taken together, our results elucidate the cooperative function of the UV-B photoreceptor UVR8 with various transcription factors in the nucleus to orchestrate the expression of specific sets of downstream genes and, ultimately, mediate plant responses to UV-B light.

Two E3 ligases antagonistically regulate the UV-B response in Arabidopsis.

Photomorphogenesis is a pivotal developmental strategy used by plants to respond to environmental light levels. During emergence from the soil and the establishment of photomorphogenesis, seedlings encounter increasing levels of UV-B irradiation and develop adaptive responses accordingly. However, the molecular mechanisms that orchestrate UV-B signaling cascades remain elusive. Here, we provide biochemical and genetic evidence that the prolonged signaling circuits of UV-B-induced photomorphogenesis involve two sets of E3 ligases and a transcription factor in Arabidopsis thaliana The UV-B-inducible protein RUP1/RUP2 associates with the CUL4-DDB1 scaffold to form an E3 ligase, which represses photomorphogenesis by mediating the degradation of HY5, the hub transcription factor in the light signaling pathway. Conversely, COP1 directly targets RUP1/RUP2 for ubiquitination and degradation, leading to balanced RUP1/RUP2 accumulation, alleviation of the COP1-HY5 interaction, and stabilization of HY5 protein. Therefore, our study reveals that these two E3-substrate modules, CUL4-DDB1-RUP1/RUP2-HY5 and COP1-RUP1/RUP2, constitute the repression and derepression machinery by which plants respond to prolonged UV-B irradiation in photomorphogenic development.

The DTH8-Hd1 Module Mediates Day-Length-Dependent Regulation of Rice Flowering.

Photoperiodic flowering is one of the most important pathways to govern flowering in rice (Oryza sativa), in which Heading date 1 (Hd1), an ortholog of the Arabidopsis CONSTANS gene, encodes a pivotal regulator. Hd1 promotes flowering under short-day conditions (SD) but represses flowering under long-day conditions (LD) by regulating the expression of Heading date 3a (Hd3a), the FLOWERING LOCUS T (FT) ortholog in rice. However, the molecular mechanism of how Hd1 changes its regulatory activity in response to day length remains largely unknown. In this study, we demonstrated that the repression of flowering in LD by Hd1 is dependent on the transcription factor DAYS TO HEADING 8 (DTH8). Loss of DTH8 function results in the activation of Hd3a by Hd1, leading to early flowering. We found that Hd1 directly interacts with DTH8 and that the formation of the DTH8-Hd1 complex is necessary for the transcriptional repression of Hd3a by Hd1 in LD, implicating that the switch of Hd1 function is mediated by DTH8 in LD rather than in SD. Furthermore, we revealed that DTH8 associates with the Hd3a promoter to modulate the level of H3K27 trimethylation (H3K27me3) at the Hd3a locus. In the presence of the DTH8-Hd1 complex, the H3K27me3 level was increased at Hd3a, whereas loss of DTH8 function resulted in decreased H3K27me3 level at Hd3a. Taken together, our findings indicate that, in response to day length, DTH8 plays a critical role in mediating the transcriptional regulation of Hd3a by Hd1 through the DTH8-Hd1 module to shape epigenetic modifications in photoperiodic flowering.

Coordinated photomorphogenic UV-B signaling network captured by mathematical modeling.

Long-wavelength and low-fluence UV-B light is an informational signal known to induce photomorphogenic development in plants. Using the model plant Arabidopsis thaliana, a variety of factors involved in UV-B-specific signaling have been experimentally characterized over the past decade, including the UV-B light receptor UV resistance locus 8; the positive regulators constitutive photomorphogenesis 1 and elongated hypocotyl 5; and the negative regulators cullin4, repressor of UV-B photomorphogenesis 1 (RUP1), and RUP2. Individual genetic and molecular studies have revealed that these proteins function in either positive or negative regulatory capacities for the sufficient and balanced transduction of photomorphogenic UV-B signal. Less is known, however, regarding how these signaling events are systematically linked. In our study, we use a systems biology approach to investigate the dynamic behaviors and correlations of multiple signaling components involved in Arabidopsis UV-B-induced photomorphogenesis. We define a mathematical representation of photomorphogenic UV-B signaling at a temporal scale. Supplemented with experimental validation, our computational modeling demonstrates the functional interaction that occurs among different protein complexes in early and prolonged response to photomorphogenic UV-B.

Beyond repression of photomorphogenesis: role switching of COP/DET/FUS in light signaling.

Light is a pivotal environmental stimulus that promotes plant photomorphogenesis. Substantial progress has been achieved in defining the central repressors of photomorphogenesis, the CONSTITUTIVE PHOTOMORPHOGENIC/DE-ETIOLATED/FUSCA (COP/DET/FUS) loci, in the past 20 years. COP/DET/FUS proteins are well-conserved, and regulate a variety of biological processes in plants and animals. The fact that these proteins contribute to the repression of plant photomorphogenesis by regulating the ubiquitin-proteasome-dependent pathway has been well established. Recently, molecular insight has been gained into the functional diversity of COP/DET/FUS. Here, we review the current research on the roles of COP/DET/FUS, with a focus on the functional conversion of COP1 in photomorphogenesis.

Photoactivated UVR8-COP1 module determines photomorphogenic UV-B signaling output in Arabidopsis.

In Arabidopsis, ultraviolet (UV)-B-induced photomorphogenesis is initiated by a unique photoreceptor UV resistance locus 8 (UVR8) which utilizes its tryptophan residues as internal chromophore to sense UV-B. As a result of UV-B light perception, the UVR8 homodimer shaped by its arginine residues undergoes a conformational switch of monomerization. Then UVR8 associates with the constitutively photomorphogenic 1-suppressor of PHYA (COP1-SPA) core complex(es) that is released from the cullin 4-damaged dna binding protein 1 (CUL4-DDB1) E3 apparatus. This association, in turn, causes COP1 to convert from a repressor to a promoter of photomorphogenesis. It is not fully understood, however, regarding the biological significance of light-absorbing and dimer-stabilizing residues for UVR8 activity in photomorphogenic UV-B signaling. Here, we take advantage of transgenic UVR8 variants to demonstrate that two light-absorbing tryptophans, W233 and W285, and two dimer-stabilizing arginines, R286 and R338, play pivotal roles in UV-B-induced photomorphogenesis. Mutation of each residue results in alterations in UV-B light perception, UVR8 monomerization and UVR8-COP1 association in response to photomorphogenic UV-B. We also identify and functionally characterize two constitutively active UVR8 variants, UVR8W285A and UVR8R338A, whose photobiological activities are enhanced by the repression of CUL4, a negative regulator in this pathway. Based on our molecular and biochemical evidence, we propose that the UVR8-COP1 affinity in plants critically determines the photomorphogenic UV-B signal transduction coupling with UVR8-mediated UV-B light perception.

Conversion from CUL4-based COP1-SPA E3 apparatus to UVR8-COP1-SPA complexes underlies a distinct biochemical function of COP1 under UV-B.

The evolutionarily conserved constitutive photomorphogenesis 1 (COP1) is a RING and WD40 protein that functions as a substrate receptor of CULLIN4-damaged DNA binding protein 1 (CUL4-DDB1)-based E3 ubiquitin ligases in both plants and animals. In Arabidopsis, COP1 is a central repressor of photomorphogenesis in the form of COP1-suppressor of PHYA (SPA) complex(es). CUL4-DDB1-COP1-SPA suppresses the photomorphogenic program by targeting the transcription factor elongated hypocotyl 5 for degradation. Intriguingly, under photomorphogenic UV-B light, COP1 reverses its repressive role and promotes photomorphogenesis. However, the mechanism by which COP1 is functionally switched is still obscure. Here, we demonstrate that UV-B triggers the physical and functional disassociation of the COP1-SPA core complex(es) from CUL4-DDB1 and the formation of a unique complex(es) containing the UV-B receptor UV resistance locus 8 (UVR8). The establishment of this UV-B-dependent COP1 complex(es) is associated with its positive modulation of elongated hypocotyl 5 stability and activity, which sheds light on the mechanism of COP1's promotive action in UV-B-induced photomorphogenesis.

Arabidopsis FHY3 and HY5 positively mediate induction of COP1 transcription in response to photomorphogenic UV-B light.

As sessile organisms, higher plants have evolved the capacity to sense and interpret diverse light signals to modulate their development. In Arabidopsis thaliana, low-intensity and long-wavelength UV-B light is perceived as an informational signal to mediate UV-B-induced photomorphogenesis. Here, we report that the multifunctional E3 ubiquitin ligase, CONSTITUTIVE PHOTOMORPHOGENESIS1 (COP1), a known key player in UV-B photomorphogenic responses, is also a UV-B-inducible gene. Two transcription factors, FAR-RED ELONGATED HYPOCOTYL3 (FHY3) and ELONGATED HYPOCOTYL5 (HY5), directly bind to distinct regulatory elements within the COP1 promoter, which are essential for the induction of the COP1 gene mediated by photomorphogenic UV-B signaling. Absence of FHY3 results in impaired UV-B-induced hypocotyl growth and reduced tolerance against damaging UV-B. Thus, FHY3 positively regulates UV-B-induced photomorphogenesis by directly activating COP1 transcription, while HY5 promotes COP1 expression via a positive feedback loop. Furthermore, FHY3 and HY5 physically interact with each other, and this interaction is diminished by UV-B. Together, our findings reveal that COP1 gene expression in response to photomorphogenic UV-B is controlled by a combinatorial regulation of FHY3 and HY5, and this UV-B-specific working mode of FHY3 and HY5 is distinct from that in far-red light and circadian conditions.