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 PIFs in various cross-talks between light, hormone and environmental cues.

Nucleotide limitation and imbalance is a well described phenomenon in animal research but understudied in the plant field. A peculiarity of pyrimidine de novo synthesis in plants is the complex subcellular organization. Here, we studied two organellar localized enzymes in the pathway, with chloroplast aspartate transcarbamoylase (ATC), and mitochondrial dihydroorotate dehydrogenase (DHODH). ATC knockdowns were most severely affected, exhibiting low levels of pyrimidine nucleotides, a low energy state, reduced photosynthetic capacity and accumulation of reactive oxygen species (ROS). Furthermore, altered leaf morphology and chloroplast ultrastructure were observed in ATC mutants. Although less affected, DHODH knockdown mutants showed impaired seed germination and altered mitochondrial ultrastructure. Thus, DHODH might not only be regulated by respiration, but vice versa exert a regulatory function on this process. Transcriptome analysis of an ATC-amiRNA line revealed massive alterations in gene expression with central metabolic pathways being downregulated and stress response and RNA related pathways being upregulated. In addition, genes involved in central carbon metabolism, intracellular transport and respiration were markedly downregulated in ATC mutants, being most likely responsible for the observed impaired growth. We conclude that impairment of the first committed step in pyrimidine metabolism, catalyzed by ATC, leads to nucleotide limitation and by this has far reaching consequences on metabolism and gene expression. DHODH might closely interact with mitochondrial respiration, as seen in delayed germination, being the reason for its localization in this organelle.

Aurones constitute one of the major classes of flavonoids, with a characteristic furanone structure that acts as the C-ring of flavonoids. Members of various enzyme families are involved in aurone biosynthesis in different higher plants, suggesting that during evolution plants acquired the ability to biosynthesize aurones independently and convergently. Bryophytes also produce aurones, but the biosynthetic pathways and enzymes involved have not been determined. The present study describes the identification and characterization of a polyphenol oxidase (PPO) that acts as an aureusidin synthase (MpAS1) in the model liverwort, Marchantia polymorpha. Crude enzyme assays using an M. polymorpha line overexpressing MpMYB14 with high accumulation of aureusidin showed that aureusidin was biosynthesized from naringenin chalcone and converted to riccionidin A. This activity was inhibited by N-phenylthiourea, an inhibitor specific to enzymes of the PPO family. Of the six PPOs highly induced in the line overexpressing MpMyb14, one, MpAS1, was found to biosynthesize aureusidin from naringenin chalcone when expressed in Saccharomyces cerevisiae. MpAS1 also recognized eriodictyol chalcone, isoliquiritigenin and butein, showing the highest activity for eriodictyol chalcone. Members of the PPO family in M. polymorpha evolved independently from PPOs in higher plants, indicating that aureusidin synthases evolved in parallel in land plants.

Hyponastic Leaves 1 (HYL1) [also known as Double-stranded RNA-Binding protein 1 (DRB1)] is a double-stranded RNA-binding protein involved in microRNA (miRNA) processing in plants. It is a core component of the Microprocessor complex and enhances the efficiency and precision of miRNA processing by the Dicer-Like 1 protein. In this work, we report a novel function of the HYL1 protein in the transcription of miRNA (MIR) genes. HYL1 colocalizes with RNA polymerase II and affects its distribution along MIR genes. Moreover, proteomic experiments revealed that the HYL1 protein interacts with many transcription factors. Finally, we show that the action of HYL1 is not limited to MIR genes and impacts the expression of many other genes, a majority of which are involved in plastid organization. These discoveries indicate HYL1 as an additional player in gene regulation at the transcriptional level, independent of its role in miRNA biogenesis.

Reactive oxygen species (ROS) are highly reactive molecules, generated by nicotinamide adenine dinucleotide phosphate oxidases encoded by respiratory burst oxidase homologs. The functions of the OsRbohs gene family in rice are diverse and poorly understood. OsRbohI was recently identified as a newly evolved gene in the rice OsRbohs gene family. However, the function of OsRbohI in regulating rice growth is not yet reported. In this study, our results indicate that knockout (KO) OsRbohI mutants showed significantly shorter shoot and primary roots, along with lower ROS content than the control lines, whereas the overexpression (OE) lines displayed contrasting results. Further experiments showed that the abnormal length of the shoot and root is mainly caused by altered cell size. These results indicate that OsRbohI regulates rice shoot and root growth through the ROS signal. More importantly, RNA-seq analysis and jasmonic acid (JA) treatment demonstrated that OsRbohI regulates rice growth via the JA synthesis and signaling pathways. Compared with the control, the results showed that the KO mutants were more sensitive to JA, whereas the OE lines were less sensitive to JA. Collectively, our results reveal a novel pathway in which OsRbohI regulates rice growth and development by affecting their ROS homeostasis through JA synthesis and signaling pathway.

In plants, microRNA (miRNA)-target interactions (MTIs) require high complementarity, a feature from which bioinformatic programs have predicted numerous and diverse targets for any given miRNA, promoting the idea of complex miRNA networks. Opposing this is a hypothesis of constrained miRNA specificity, in which functional MTIs are restricted to the few targets whose required expression output is compatible with the expression of the miRNA. To explore these opposing views, the bioinformatic pipeline Targets Ranked Using Experimental Evidence was applied to strongly conserved miRNAs to identity their high-evidence (HE) targets across species. For each miRNA family, HE targets predominantly consisted of homologs from one conserved target gene family (primary family). These primary families corresponded to the known canonical miRNA-target families, validating the approach. Very few additional HE target families were identified (secondary family), and if so, they were likely functionally related to the primary family. Many primary target families contained highly conserved nucleotide sequences flanking their miRNA-binding sites that were enriched in HE homologs across species. A number of these flanking sequences are predicted to form conserved RNA secondary structures that preferentially base pair with the miRNA-binding site, implying that these sites are highly structured. Our findings support a target landscape view that is dominated by the conserved primary target families, with a minority of either secondary target families or non-conserved targets. This is consistent with the constrained hypothesis of functional miRNA specificity, which potentially in part is being facilitated by features beyond complementarity.

In addition to being important primary productive forces in marine ecosystems, diatoms are also rich in bioactive substances such as triacylglycerol and fucoxanthin. However, little is known about the transcriptional mechanisms underlying the biosynthesis of these substances. In this study, we found that the heat shock transcription factor PtHSF1 positively regulated the synthesis of triacylglycerol and fucoxanthin in Phaeodactylum tricornutum. Overexpression of PtHSF1 could increase the contents of triacylglycerol and fucoxanthin and upregulate key enzyme genes involved in the triacylglycerol and fucoxanthin biosynthesis pathways. On the other hand, gene silencing of PtHSF1 reduced the contents of triacylglycerol and fucoxanthin and the expression of the key enzyme genes involved in the triacylglycerol and fucoxanthin biosynthesis pathways. Further biochemical analysis revealed that PtHSF1 upregulated glycerol-2-phosphate acyltransferase 3 (GPAT3) and 1-deoxy-d-xylulose 5-phosphate synthase (DXS) by directly binding to their promoters, while genetic analysis demonstrated that PtHSF1 acted upstream of GPAT3 and DXS to regulate triacylglycerol and fucoxanthin synthesis. Therefore, in addition to elucidating the regulation mechanisms underlying PtHSF1-mediated triacylglycerol and fucoxanthin synthesis, this study also provided a candidate target for metabolic engineering of triacylglycerol and fucoxanthin in P. tricornutum.

Unicellular diazotrophic cyanobacteria contribute significantly to the photosynthetic productivity of the ocean and the fixation of molecular nitrogen, with photosynthesis occurring during the day and nitrogen fixation during the night. In species like Crocosphaera watsonii WH8501, the decline in photosynthetic activity in the night is accompanied by the disassembly of oxygen-evolving photosystem II (PSII) complexes. Moreover, in the second half of the night phase, a small amount of rogue D1 (rD1), which is related to the standard form of the D1 subunit found in oxygen-evolving PSII, but of unknown function, accumulates but is quickly degraded at the start of the light phase. We show here that the removal of rD1 is independent of the rD1 transcript level, thylakoid redox state and trans-thylakoid pH but requires light and active protein synthesis. We also found that the maximal level of rD1 positively correlates with the maximal level of chlorophyll (Chl) biosynthesis precursors and enzymes, which suggests a possible role for rogue PSII (rPSII) in the activation of Chl biosynthesis just before or upon the onset of light, when new photosystems are synthesized. By studying strains of Synechocystis PCC 6803 expressing Crocosphaera rD1, we found that the accumulation of rD1 is controlled by the light-dependent synthesis of the standard D1 protein, which triggers the fast FtsH2-dependent degradation of rD1. Affinity purification of FLAG-tagged rD1 unequivocally demonstrated the incorporation of rD1 into a non-oxygen-evolving PSII complex, which we term rPSII. The complex lacks the extrinsic proteins stabilizing the oxygen-evolving Mn4CaO5 cluster but contains the Psb27 and Psb28-1 assembly factors.

Seed dormancy and germination are regulated by endogenous gene expression as well as hormonal and environmental conditions, such as salinity, which greatly inhibits seed germination. MOTHER OF FT AND TFL1 (MFT), which encodes a phosphatidylethanolamine-binding protein, is a key regulator of seed germination in Arabidopsis thaliana. There are two orthologous genes of AtMFT in rice (Oryza sativa), namely, OsMFT1 and OsMFT2. However, the functions of these two genes in regulating rice seed germination under salt stress remain unknown. In this study, we found that seeds of loss-of-function osmft1 mutants germinated faster than wild-type (WT) seeds under salt stress, but this was not the case for loss-of-function osmft2 mutants. Overexpression of OsMFT1 (OsMFT1OE) or OsMFT2 increased the sensitivity to salt stress during seed germination. Transcriptome comparisons of osmft1 vs WT in the absence and presence of salt stress yielded several differentially expressed genes, which were associated with salt stress, plant hormone metabolism and signaling pathways, such as B-BOX ZINC FINGER 6, O. sativa bZIP PROTEIN 8 and GIBBERELLIN (GA) 20-oxidase 1. In addition, the sensitivity of OsMFT1OE seeds to GA and osmft1 seeds to abscisic acid (ABA) during seed germination increased under salt stress. Overall, our results indicate that ABA and GA metabolism and their signaling pathways are regulated by OsMFT1, modulating seed germination in rice under salt stress.

The chloroplast signal recognition particle (CpSRP) receptor (CpFTSY) is a component of the CpSRP pathway that post-translationally targets light-harvesting complex proteins (LHCPs) to the thylakoid membranes in plants and green algae containing chloroplasts derived from primary endosymbiosis. In plants, CpFTSY also plays a major role in the co-translational incorporation of chloroplast-encoded subunits of photosynthetic complexes into the thylakoids. This role has not been demonstrated in green algae. So far, its function in organisms with chloroplasts derived from secondary endosymbiotic events has not been elucidated. Here, we report the generation and characterization of mutants lacking CpFTSY in the diatom Phaeodactylum tricornutum. We found that this protein is not involved in inserting LHCPs into thylakoid membranes, indicating that the post-translational part of the CpSRP pathway is not active in this group of microalgae. The lack of CpFTSY caused an increased level of photoprotection, low electron transport rates, inefficient repair of photosystem II (PSII), reduced growth, a strong decline in the PSI subunit PsaC and upregulation of proteins that might compensate for a non-functional co-translational CpSRP pathway during light stress conditions. The phenotype was highly similar to the one described for diatoms lacking another component of the co-translational CpSRP pathway, the CpSRP54 protein. However, in contrast to cpsrp54 mutants, only one thylakoid membrane protein, PetD of the Cytb6f complex, was downregulated in cpftsy. Our results point to a minor role for CpFTSY in the co-translational CpSRP pathway, suggesting that other mechanisms may partially compensate for the effect of a disrupted CpSRP pathway.

Hypocotyl elongation is inhibited by light and promoted by darkness. The plant hormone abscisic acid (ABA) also inhibits hypocotyl elongation. However, details of the molecular mechanism that regulates the integrated effects of light and ABA signaling on hypocotyl elongation remain unclear. Long non-coding RNAs (lncRNAs; >200 nt) do not encode proteins but play many physiological roles in organisms. Until now, only a few lncRNAs related to hypocotyl elongation have been reported. The lncRNAs BoNR8 (272 nt) and AtR8 (259 nt), both of which are transcribed by RNA polymerase III, are homologous lncRNAs that are abundantly present in cabbage and Arabidopsis, respectively. These lncRNAs shared 77% sequence identity, and their predicted RNA secondary structures were similar; the non-conserved nucleotides in both sequences were positioned mainly in the stem-loop regions of the secondary structures. A previous study showed that BoNR8 regulated seed germination along with ABA and that AtR8 may be involved in innate immune function in Arabidopsis. Our results show that the expression levels of BoNR8 and AtR8 were differentially affected by light and ABA and that overexpression (OX) of both BoNR8 and AtR8 in Arabidopsis regulated hypocotyl elongation depending on light and ABA.. The expression levels of light-related genes PHYB, COP1, HY5 and PIF4 and ABA-related genes ABI3 and ABI5 were altered in the AtR8-OX and BoNR8-OX lines, and, in an ABI3-defective mutant, hypocotyl elongation was greatly increased under dark condition with the addition of ABA. These results indicate that BoNR8 and AtR8 regulate hypocotyl elongation together with ABI3 and key downstream light signaling genes.

The huge structural diversity exhibited by plant specialized metabolites has primarily been considered to result from the catalytic specificity of their biosynthetic enzymes. Accordingly, enzyme gene multiplication and functional differentiation through spontaneous mutations have been established as the molecular mechanisms that drive metabolic evolution. Nevertheless, how plants have assembled and maintained such metabolic enzyme genes and the typical clusters that are observed in plant genomes, as well as why identical specialized metabolites often exist in phylogenetically remote lineages, are currently explained only poorly by a concept known as convergent evolution. Here, we compile recent knowledge on the co-presence of metabolic modules that are common in the plant kingdom but have evolved under specific historical and contextual constraints defined by the physicochemical properties of each plant specialized metabolite and the genetic presets of the biosynthetic genes. Further, we discuss a common manner to generate uncommon metabolites (uniqueness out of uniformity) and an uncommon manner to generate common metabolites (uniqueness behind uniformity). This review describes the emerging aspects of the evolvability of plant specialized metabolism that underlie the vast structural diversity of plant specialized metabolites in nature.