SlCPK27 cross-links SlHY5 and SlPIF4 in brassinosteroid-dependent photo- and thermo-morphogenesis in tomato.

ELONGATED HYPOCOTOYL5 (HY5) and PHYTOCHROME INTERACTING FACTORs (PIFs) are two types of important light-related regulators of plant growth, however, their interplay remains elusive. Here, we report that the activated tomato (Solanum lycopersicum) HY5 (SlHY5) triggers the transcription of a Calcium-dependent Protein Kinase SlCPK27. SlCPK27 interacts with and phosphorylates SlPIF4 at Ser-252 and Ser-308 phosphosites to promote its degradation. SlPIF4 promotes hypocotyl elongation mainly by activating the transcription of SlDWF, a key gene in brassinosteroid (BR) biosynthesis. Such a SlHY5-SlCPK27-SlPIF4-BR cascade not only plays a crucial role in photomorphogenesis but also regulates thermomorphogenesis. Our results uncover a previously unidentified mechanism that integrates Ca2+ signaling with the light signaling pathways to regulate plant growth by modulating BR biosynthesis in response to changes in ambient light and temperature.

Phase Separation of Phytochrome B in HEK293T Cells.

In this method, we employed HEK293T cells to express the plant photoreceptor phytochrome B (phyB). Through the application of various treatments such as phycocyanobilin (PCB) supplementation, red light exposure, and temperature adjustments, the phyB proteins exhibited liquid-liquid phase separation, leading to the formation of biomolecular condensates. Here, we present a comprehensive description of the protein expression, cell treatment, and imaging capture procedures. This detailed guide provides step-by-step instructions on how to induce phase separation of phyB proteins in HEK293T cells. By utilizing this approach, researchers can investigate the physicochemical characteristics and dynamic formation process of phyB photobodies with precision.

Modulation of starch synthesis in Arabidopsis via phytochrome B-mediated light signal transduction.

Starch is a major storage carbohydrate in plants and is critical in crop yield and quality. Starch synthesis is intricately regulated by internal metabolic processes and external environmental cues; however, the precise molecular mechanisms governing this process remain largely unknown. In this study, we revealed that high red to far-red (high R:FR) light significantly induces the synthesis of leaf starch and the expression of synthesis-related genes, whereas low R:FR light suppress these processes. Arabidopsis phytochrome B (phyB), the primary R and FR photoreceptor, was identified as a critical positive regulator in this process. Downstream of phyB, basic leucine zipper transcription factor ELONGATED HYPOCOTYL5 (HY5) was found to enhance starch synthesis, whereas the basic helix-loop-helix transcription factors PHYTOCHROME INTERACTING FACTORs (PIF3, PIF4, and PIF5) inhibit starch synthesis in Arabidopsis leaves. Notably, HY5 and PIFs directly compete for binding to a shared G-box cis-element in the promoter region of genes encoding starch synthases GBSS, SS3, and SS4, which leads to antagonistic regulation of their expression and, consequently, starch synthesis. Our findings highlight the vital role of phyB in enhancing starch synthesis by stabilizing HY5 and facilitating PIFs degradation under high R:FR light conditions. Conversely, under low R:FR light, PIFs predominantly inhibit starch synthesis. This study provides insight into the physiological and molecular functions of phyB and its downstream transcription factors HY5 and PIFs in starch synthesis regulation, shedding light on the regulatory mechanism by which plants synchronize dynamic light signals with metabolic cues to module starch synthesis.

Application of Multi-Omics Technologies to the Study of Phytochromes in Plants.

Phytochromes (phy) are distributed in various plant organs, and their physiological effects influence plant germination, flowering, fruiting, and senescence, as well as regulate morphogenesis throughout the plant life cycle. Reactive oxygen species (ROS) are a key regulatory factor in plant systemic responses to environmental stimuli, with an attractive regulatory relationship with phytochromes. With the development of high-throughput sequencing technology, omics techniques have become powerful tools, and researchers have used omics techniques to facilitate the big data revolution. For an in-depth analysis of phytochrome-mediated signaling pathways, integrated multi-omics (transcriptomics, proteomics, and metabolomics) approaches may provide the answer from a global perspective. This article comprehensively elaborates on applying multi-omics techniques in studying phytochromes. We describe the current research status and future directions on transcriptome-, proteome-, and metabolome-related network components mediated by phytochromes when cells are subjected to various stimulation. We emphasize the importance of multi-omics technologies in exploring the effects of phytochromes on cells and their molecular mechanisms. Additionally, we provide methods and ideas for future crop improvement.

Temporal control of the Aux/IAA genes BnIAA32 and BnIAA34 mediates Brassica napus dual shade responses.

Precise responses to changes in light quality are crucial for plant growth and development. For example, hypocotyls of shade-avoiding plants typically elongate under shade conditions. Although this typical shade-avoidance response (TSR) has been studied in Arabidopsis (Arabidopsis thaliana), the molecular mechanisms underlying shade tolerance are poorly understood. Here we report that B. napus (Brassica napus) seedlings exhibit dual shade responses. In addition to the TSR, B. napus seedlings also display an atypical shade response (ASR), with shorter hypocotyls upon perception of early-shade cues. Genome-wide selective sweep analysis indicated that ASR is associated with light and auxin signaling. Moreover, genetic studies demonstrated that phytochrome A (BnphyA) promotes ASR, whereas BnphyB inhibits it. During ASR, YUCCA8 expression is activated by early-shade cues, leading to increased auxin biosynthesis. This inhibits hypocotyl elongation, as young B. napus seedlings are highly sensitive to auxin. Notably, two non-canonical AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) repressor genes, BnIAA32 and BnIAA34, are expressed during this early stage. BnIAA32 and BnIAA34 inhibit hypocotyl elongation under shade conditions, and mutations in BnIAA32 and BnIAA34 suppress ASR. Collectively, our study demonstrates that the temporal expression of BnIAA32 and BnIAA34 determines the behavior of B. napus seedlings following shade-induced auxin biosynthesis.

Combined Transcriptomic and Metabolomic Approach Revealed a Relationship between Light Control, Photoprotective Pigments, and Lipid Biosynthesis in Olives.

Olive possesses excellent nutritional and economic values for its main healthy products. Among them, a high content of antioxidant compounds, balanced during the ripening process, are produced under genetic and environmental control, resulting in high variability among cultivars. The genes involved in these complex pathways are mainly known, but despite many studies which indicated the key role of light quality and quantity for the synthesis of many metabolites in plants, limited information on these topics is available in olive. We carried out a targeted gene expression profiling in three olive cultivars, Cellina di Nardò, Ruveia, and Salella, which were selected for their contrasting oleic acid and phenolic content. The -omics combined approach revealed a direct correlation between a higher expression of the main flavonoid genes and the high content of these metabolites in 'Cellina di Nardò'. Furthermore, it confirmed the key role of FAD2-2 in the linoleic acid biosynthesis. More interestingly, in all the comparisons, a co-regulation of genes involved in photoperception and circadian clock machinery suggests a key role of light in orchestrating the regulation of these pathways in olive. Therefore, the identified genes in our analyses might represent a useful tool to support olive breeding, although further investigations are needed.

Light response of gametophyte in Adiantum flabellulatum: transcriptome analysis and identification of key genes and pathways.

Light serves not only as a signaling cue perceived by plant photoreceptors but also as an essential energy source captured by chloroplasts. However, excessive light can impose stress on plants. Fern gametophytes possess the unique ability to survive independently and play a critical role in the alternation of generations. Due to their predominantly shaded distribution under canopies, light availability becomes a limiting factor for gametophyte survival, making it imperative to investigate their response to light. Previous research on fern gametophytes' light response has been limited to the physiological level. In this study, we examined the light response of Adiantum flabellulatum gametophytes under different photosynthetic photon flux density (PPFD) levels and identified their high sensitivity to low light. We thereby determined optimal and stress-inducing light conditions. By employing transcriptome sequencing, weighted gene co-expression network analysis, and Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses, we identified 10,995 differentially expressed genes (DEGs). Notably, 3 PHYBs and 5 Type 1 CRYs (CRY1s) were significantly down-regulated at low PPFD (0.1 µmol m-2 s-1). Furthermore, we annotated 927 DEGs to pathways related to photosynthesis and 210 to the flavonoid biosynthesis pathway involved in photoprotection. Additionally, we predicted 34 transcription factor families and identified a close correlation between mTERFs and photosynthesis, as well as a strong co-expression relationship between MYBs and bHLHs and genes encoding flavonoid synthesis enzymes. This comprehensive analysis enhances our understanding of the light response of fern gametophytes and provides novel insights into the mechanisms governing their responses to light.

Emerging Roles of FHY3 and FAR1 as System Integrators in Plant Development.

FAR-RED ELONGATED HYPOCOTYL3 (FHY3) and its homolog FAR-RED-IMPAIRED RESPONSE1 (FAR1) are transcription factors derived from transposases essential for phytochrome A-mediated light signaling. In addition to their essential role in light signaling, FHY3 and FAR1 also play diverse regulatory roles in plant growth and development, including clock entrainment, seed dormancy and germination, senescence, chloroplast formation, branching, flowering and meristem development. Notably, accumulating evidence indicates that the emerging role of FHY3 and FAR1 in environmental stress signaling has begun to be revealed. In this review, we summarize these recent findings in the context of FHY3 and FAR1 as integrators of light and other developmental and stressful signals. We also discuss the antagonistic action of FHY3/FAR1 and Phytochrome Interating Factors (PIFs) in various cross-talks between light, hormone and environmental cues.

Genome resequencing and transcriptome analysis reveal the molecular mechanism of albinism in Cordyceps militaris.

Light is an important regulator of most fungal life activities and transmits signals through certain photoreceptor proteins such as phytochromes and cryptochromes. However, the light response mechanism varies across different fungi. The WCC complex composed of white collar-1 (WC-1) and white collar-2 (WC-2) is considered to be the key factor regulating fungal albinism. The photoreceptor protein Vivid (VVD) is the negative regulator of the WCC complex. In this study, we discovered an albino mutant (Alb) generated by 60Co-γ-ray irradiation from Cordyceps militaris (C. militaris). This mutant showed albinism of the mycelia and fruiting bodies under light, and the fruiting bodies developed normally. However, this phenotype in Alb differed from that in the CmWC-1 mutant. This suggests that CmWC1 may not be mutated in Alb. A mutated polyketide synthase (CmPKS) was found through genome resequencing analysis. CmPKS was significantly induced by a light signal, and its mutation reduced melanin accumulation in C. militaris. In addition, we found that a zinc-finger domain-containing protein (CmWC-3) was induced by a light signal and interacted with CmWC-1 and CmVVD. Moreover, CmWC-2 also interacted with CmWC-1 to form the WCC complex and was inhibited by CmVVD. In addition, CmWC-3 directly bound with the CmPKS promoter, but CmWC1 did not. These results suggest that albinism and fruiting body development are two independent processes; the WCC complex of CmWC-1 with CmWC-3 regulates CmPKS expression to regulate color change, whereas CmWC-1 with CmWC-2 affects fruiting body development via the carotenoid pathway. These findings will help us to better understand the albinism mechanism of C. militaris.

HY5 functions as a systemic signal by integrating BRC1-dependent hormone signaling in tomato bud outgrowth.

Light plays an important role in determining plant architecture, which greatly influences crop yield. However, the precise mechanisms by which light signaling regulates bud outgrowth remain to be identified. Here, we show that light regulates bud outgrowth via both HY5 and brassinosteroid (BR)-dependent pathways in tomato. Inactivation of the red-light photoreceptor PHYB, or deficiencies in PHYB or the blue-light photoreceptor CRY1a, inhibits bud outgrowth and leads to decreased accumulation of HY5 protein and increased transcript level of BRANCHED1 (BRC1), a central integrator of branching signals. HY5, functioning as a mobile systemic signal from leaves, promotes bud outgrowth by directly suppressing BRC1 transcript and activating the transcript of BR biosynthesis genes within the lateral buds in tomato. Furthermore, BRC1 prevents the accumulation of cytokinin (CK) and gibberellin (GA) by directly inhibiting the transcript of CK synthesis gene LOG4, while increasing the transcript levels of CK and GA degradation genes (CKX7, GA2ox4, and GA2ox5), leading to an arrest of bud outgrowth. Moreover, bud outgrowth occurs predominantly in the day due to the suppression of BRC1 transcript by HY5. These findings demonstrate that light-inducible HY5 acts as a systemic signaling factor in fine-tuning the bud outgrowth of tomato.

The DELLA-ABI4-HY5 module integrates light and gibberellin signals to regulate hypocotyl elongation.

Plant growth is coordinately controlled by various environmental and hormonal signals, of which light and gibberellin (GA) signals are two critical factors with opposite effects on hypocotyl elongation. Although interactions between the light and GA signaling pathways have been studied extensively, the detailed regulatory mechanism of their direct crosstalk in hypocotyl elongation remains to be fully clarified. Previously, we reported that ABA INSENSITIVE 4 (ABI4) controls hypocotyl elongation through its regulation of cell-elongation-related genes, but whether it is also involved in GA signaling to promote hypocotyl elongation is unknown. In this study, we show that promotion of hypocotyl elongation by GA is dependent on ABI4 activation. DELLAs interact directly with ABI4 and inhibit its DNA-binding activity. In turn, ABI4 combined with ELONGATED HYPOCOTYL 5 (HY5), a key positive factor in light signaling, feedback regulates the expression of the GA2ox GA catabolism genes and thus modulates GA levels. Taken together, our results suggest that the DELLA-ABI4-HY5 module may serve as a molecular link that integrates GA and light signals to control hypocotyl elongation.

Sensory circuitry controls cytosolic calcium-mediated phytochrome B phototransduction.

Although Ca2+ has long been recognized as an obligatory intermediate in visual transduction, its role in plant phototransduction remains elusive. Here, we report a Ca2+ signaling that controls photoreceptor phyB nuclear translocation in etiolated seedlings during dark-to-light transition. Red light stimulates acute cytosolic Ca2+ increases via phyB, which are sensed by Ca2+-binding protein kinases, CPK6 and CPK12 (CPK6/12). Upon Ca2+ activation, CPK6/12 in turn directly interact with and phosphorylate photo-activated phyB at Ser80/Ser106 to initiate phyB nuclear import. Non-phosphorylatable mutation, phyBS80A/S106A, abolishes nuclear translocation and fails to complement phyB mutant, which is fully restored by combining phyBS80A/S106A with a nuclear localization signal. We further show that CPK6/12 function specifically in the early phyB-mediated cotyledon expansion, while Ser80/Ser106 phosphorylation generally governs phyB nuclear translocation. Our results uncover a biochemical regulatory loop centered in phyB phototransduction and provide a paradigm for linking ubiquitous Ca2+ increases to specific responses in sensory stimulus processing.

l-Tryptophan synergistically increased carotenoid accumulation with blue light in maize (Zea mays L.) sprouts.

In the present study, l-tryptophan was applied in combination with blue light to modulate carotenoid biosynthesis in maize sprouts. The profiles of carotenoids, chlorophylls, and relative genes in carotenoid biosynthesis and light signaling pathways were studied. l-tryptophan and blue light both promoted the accumulation of carotenoids, and their combination further increased carotenoid content by 120%. l-tryptophan exerted auxin-like effects and stimulated PSY expression in blue light exposure maize sprouts, resulting in increased α- and ß- carotenes. l-tryptophan could also play a photoprotective role through the xanthophyll cycle under blue light. In addition, CRY in the light signaling pathway was critical for carotenoid biosynthesis. These findings provide new insights into the regulation of carotenoid biosynthesis and l-tryptophan could be used in conjunction with blue light to fortify carotenoids in maize sprouts.

Effect of light quality on polyphenol biosynthesis in three varieties of mung bean sprouts with different color seed coats.

KEY MESSAGE: We investigated the mechanism of the effect of different light qualities on the synthesis and regulation of mung bean sprouts. Light quality acts as a signal molecule, strongly enhancing polyphenol biosynthesis in sprouts. Mung bean (Vigna radiata) sprouts are a popular sprouting vegetable all over the world and are an excellent source of polyphenols with high antioxidant activity. This study investigated the effects of light qualities on the kinetic changes and metabolic regulation mechanism of light signal-mediating polyphenols in three mung bean sprout cultivars. Experimental results showed that three light qualities significantly enhanced the contents of caffeic acid, rutin, vitexin, genistin and delphinidin 3-glucoside. Interestingly, ferulic acid and vitexin responded selectively to blue light and red light, severally. Most genes involved in polyphenol biosynthesis were activated under different light quality conditions, resulting in an overaccumulation of phenylpropanoids. Pearson correlation analysis showed that PAL, F3H, F3'H and F3'5'H expression correlated highly with rutin, whereas ANS expression paralleled anthocyanin biosynthesis. Moreover, MYB111, MYB3, MYB4, MYB1 and MYC2 were critical regulators of polyphenol biosynthesis in mung bean sprouts. These changes were likely due to the changes in the expression of the photoreceptor genes CRY-D, PHOT2, PHYE and light response genes (PIF3 and HY5). Our results provide insights into polyphenol biosynthesis in sprouts and microgreens.

AaMYB108 is the core factor integrating light and jasmonic acid signaling to regulate artemisinin biosynthesis in Artemisia annua.

Artemisinin, a sesquiterpene compound synthesized and stored in the glandular trichome of Artemisia annua leaves, has been used to treat malaria. Previous studies have shown that both light and jasmonic acid (JA) can promote the biosynthesis of artemisinin, and the promotion of artemisinin by JA is dependent on light. However, the specific molecular mechanism remains unclear. Here, we report a MYB transcription factor, AaMYB108, identified from transcriptome analysis of light and JA treatment, as a positive regulator of artemisinin biosynthesis in A. annua. AaMYB108 promotes artemisinin biosynthesis by interacting with a previously characterized positive regulator of artemisinin, AaGSW1. Then, we found that AaMYB108 interacted with AaCOP1 and AaJAZ8, respectively. The function of AaMYB108 was influenced by AaCOP1 and AaJAZ8. Through the treatment of AaMYB108 transgenic plants with light and JA, it was found that the promotion of artemisinin by light and JA depends on the presence of AaMYB108. Taken together, our results reveal the molecular mechanism of JA regulating artemisinin biosynthesis depending on light in A. annua. This study provides new insights into the integration of light and phytohormone signaling to regulate terpene biosynthesis in plants.

Bagging Strategy and Identification of Coloring Mode of 'Xinqihong' Pear.

'Xinqihong' is a recently selected and well-colored red pear (Pyrus bretschneideri Rehd.) cultivar that is popular in the marketplace owing to the bright red color and high quality of the fruit. The red pigmentation is strongly associated with the light signal. However, its responses to bagging treatment and to light exposure after shading are unknown. In this study, the fruit were treated with three types of fruit bags. 'Xinqihong' fruit colored rapidly in response to light stimulation. A white fruit bag was optimal for bagging of 'Xinqihong' fruit. To ensure satisfactory red pigmentation, the fruit required exposure to 30 days of light after bag removal. A transcriptome analysis was conducted to screen light-signal-related genes and identify their possible functions. PbCRY1 activated the promoter of PbHY5.2 and enhanced its expression. PbHY5.2 activated the promoter activity of PbUFGT and induced anthocyanin synthesis, and also showed self-activation characteristics. Both PbCRY2 and PbPHY1 induced anthocyanin accumulation. Thus, blue-light receptors played an important role in anthocyanin synthesis. This study provides a theoretical basis for the bagging cultivation of new varieties of 'Xinqihong', and lays a foundation for the study of the mechanisms of red pear fruit coloring in response to light signals.

Transcriptome, metabolome and suppressor analysis reveal an essential role for the ubiquitin-proteasome system in seedling chloroplast development.

BACKGROUND: Many regulatory circuits in plants contain steps of targeted proteolysis, with the ubiquitin proteasome system (UPS) as the mediator of these proteolytic events. In order to decrease ubiquitin-dependent proteolysis, we inducibly expressed a ubiquitin variant with Arg at position 48 instead of Lys (ubK48R). This variant acts as an inhibitor of proteolysis via the UPS, and allowed us to uncover processes that are particularly sensitive to UPS perturbation. RESULTS: Expression of ubK48R during germination leads to seedling death. We analyzed the seedling transcriptome, proteome and metabolome 24 h post ubK48R induction and confirmed defects in chloroplast development. We found that mutations in single genes can suppress seedling lethality, indicating that a single process in seedlings is critically sensitive to decreased performance of the UPS. Suppressor mutations in phototropin 2 (PHOT2) suggest that a contribution of PHOT2 to chloroplast protection is compromised by proteolysis inhibition. CONCLUSIONS: Overall, the results reveal protein turnover as an integral part of a signal transduction chain that protects chloroplasts during development.

Self-Powered Sb2S3 Thin-Film Photodetectors with High Detectivity for Weak Light Signal Detection.

Photodetectors (PDs) for weak light signal detection have wide applications for optical communication and imaging. Antimony sulfide (Sb2S3) as a nontoxic and stable light-sensitive material becomes a promising candidate for weak light PDs, which are developing in the direction of high response, high speed, and low cost. Herein, a self-powered Sb2S3 PD with the structure of FTO/TiO2/Sb2S3/Au is developed to achieve weak light detection for 300-750 nm visible light. We control the Sb2S3 thickness with about 460 nm to match depletion region width (438 nm) and obtain an excellent photoresponsivity and 3 dB bandwidth. Furtherly, we prepare pyramid structure polydimethylsiloxane (PDMS) on the illuminating surface to enhance the performance of weak light detection by light-trapping effects. The photocurrent of Sb2S3 PD with 20 µm-sized PDMS texture achieves 13.6% improvement compared with the control one. Under weak 530 nm light illumination of 1 µW cm-2, the self-powered Sb2S3 PD with PDMS achieves high responsivity (3.41 A W-1), large detectivity (2.84 × 1013 Jones), and ultrafast speed (15 µs). The present Sb2S3 PD and light-trapping strategy are expected to provide an alternative to future commercial weak light detection applications.

Carbon Monoxide (CO) as a Retinal Regulator of Heme Oxygenases -1, and -2 (HO's) Expression.

Carbon monoxide (CO) has been proposed as a chemical light signal and neural system modulator via heme oxygenases -1 and -2 (HO-1 and HO-2). Many papers have proven the CO-HO circuit to be important for such physiological pathways as the molecular biological clock and the GnRH axis, but also in such pathological occurrences as ischemic injuries, or inflammation as a regenerative and neuroprotective factor. In this in vivo experiment, we used three groups of pigs: control-housed in natural conditions without any procedures; without CO-adapted and kept in constant darkness, infused with blank plasma; and with CO-adapted and kept in constant darkness infused with CO-enriched plasma. After the experiments, each animal was slaughtered and its eyes were collected for further analysis. Quantitative PCR and Western blot analysis were performed to show statistical differences in the expressions between the experimental groups. Our data revealed that exogenous CO is regulator of mRNA transcription for HO-1 and HO-2 and PCNA. Moreover, the mRNA abundance of analyzed factors in the experimental group after CO elevation revealed a restored gene-expression level similar to the control group, which we had observed in the group's restored protein level after CO elevation. In conclusion, exogenous CO regulates HO's and PCNA gene expression on transcriptional and translational levels in a similar way as a light cue.

HY5 inhibits in vitro shoot stem cell niches initiation via directly repressing pluripotency and cytokinin pathways.

The efficiency of plant regeneration from explants is influenced by phytohormones and environmental conditions. Light has a particularly marked effect on in vitro shoot regeneration, and some light signaling factors are involved in shoot regeneration, while the underlying molecular mechanism remains elusive. Here, ELONGATED HYPOCOTYL5 (HY5), as the key transcription factor of light signaling, was found to inhibit shoot regeneration under a range of light conditions. The heightened shoot regeneration capacity of the hy5-215 mutant was less marked in the dark than in the light, showing that HY5-mediated inhibition of shoot regeneration is partly light dependent. The co-localization of WUSCHEL (WUS) and CLAVATA3 (CLV3) expressions was found to coincide with the initiation of stem cell niches in root explants during shoot regeneration. HY5 could directly repress CLV3 and WUS expression by binding to their respective promoters. In parallel, HY5 indirectly repressed CLV3 and WUS by binding to the ARABIDOPSIS RESPONSE REGULATOR12 (ARR12) promoter. The resulting dual regulation exerted by HY5 on WUS and CLV3 impeded the initiation of shoot stem cell niches. A HY5-mediated inhibitory pathway was identified that links cytokinin signaling and the pluripotency pathway during shoot regeneration.