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As one of the most important crops in the world, rice (Oryza sativa) is a model plant for metabolome research. Although many studies have focused on the analysis of specific tissues, the changes in metabolite abundance across the entire life cycle have not yet been determined. In this study, combining both targeted and nontargeted metabolite profiling methods, a total of 825 annotated metabolites were quantified in rice samples from different tissues covering the entire life cycle. The contents of metabolites in different tissues of rice were significantly different, with various metabolites accumulating in the plumule and radicle during seed germination. Combining these data with transcriptome data obtained from the same time period, we constructed the Rice Metabolic Regulation Network. The metabolites and co-expressed genes were further divided into 12 clusters according to their accumulation patterns, with members within each cluster displaying a uniform and clear pattern of abundance across development. Using this dataset, we established a comprehensive metabolic profile of the rice life cycle and used two independent strategies to identify novel transcription factors—namely the use of known regulatory genes as bait to screen for new networks underlying lignin metabolism and the unbiased identification of new glycerophospholipid metabolism regulators on the basis of tissue specificity. This study thus demonstrates how guilt-by-association analysis of metabolome and transcriptome data spanning the entire life cycle in cereal crops provides novel resources and tools to aid in understanding the mechanisms underlying important agronomic traits.

Although natural variations in rice flavonoids exist, and biochemical characterization of a few flavonoid glycosyltransferases has been reported, few studies focused on natural variations in tricin-lignan-glycosides and their underlying genetic basis. In this study, we carried out metabolic profiling of tricin-lignan-glycosides and identified a major quantitative gene annotated as a UDP-dependent glycosyltransferase OsUGT706C2 by metabolite-based genome-wide association analysis. The putative flavonoid glycosyltransferase OsUGT706C2 was characterized as a flavonoid 7-O-glycosyltransferas in vitro and in vivo. Although the in vitro enzyme activity of OsUGT706C2 was similar to that of OsUGT706D1, the expression pattern and induced expression profile of OsUGT706C2 were very different from those of OsUGT706D1. Besides, OsUGT706C2 was specifically induced by UV-B. Constitutive expression of OsUGT706C2 in rice may modulate phenylpropanoid metabolism at both the transcript and metabolite levels. Furthermore, overexpressing OsUGT706C2 can enhance UV-B tolerance by promoting ROS scavenging in rice. Our findings might make it possible to use the glycosyltransferase OsUGT706C2 for crop improvement with respect to UV-B adaptation and/or flavonoid accumulation, which may contribute to stable yield.

Plants produce specialized metabolites to adapt to the ever-changing environments. Flavonoids are antioxidants essential for growth, development, and breeding with increased stress resistance in crops. However, the mechanism of the involvement of flavonoids in ultraviolet-B (UV-B) stress in rice (Oryza sativa) is largely unknown. In this study, we cloned and functionally identified a receptor-like kinase (OsRLCK160) and a bZIP transcription factor (OsbZIP48) positively regulating flavonoid accumulation through metabolite-based genome-wide association study of the flavonoid content in rice. Meanwhile, OsRLCK160 interacted with and phosphorylated OsbZIP48 to regulate the flavonoid accumulation and participate in UV-B tolerance in rice. Our study indicates the importance of applying OsRLCK160 and OsbZIP48 to advance the fundamental understanding of stable rice production and breed UV-B-tolerant rice varieties, which may contribute to breeding high-yield rice varieties.

Diterpenoids are the major group of antimicrobial phytoalexins in rice1,2. Here, we report the discovery of a rice diterpenoid gene cluster on chromosome 7 (DGC7) encoding the entire biosynthetic pathway to 5,10-diketo-casbene, a member of the monocyclic casbene-derived diterpenoids. We revealed that DGC7 is regulated directly by JMJ705 through methyl jasmonate-mediated epigenetic control3. Functional characterization of pathway genes revealed OsCYP71Z21 to encode a casbene C10 oxidase, sought after for the biosynthesis of an array of medicinally important diterpenoids. We further show that DGC7 arose relatively recently in the Oryza genus, and that it was partly formed in Oryza rufipogon and positively selected for in japonica during domestication. Casbene-synthesizing enzymes that are functionally equivalent to OsTPS28 are present in several species of Euphorbiaceae but gene tree analysis shows that these and other casbene-modifying enzymes have evolved independently. As such, combining casbene-modifying enzymes from these different families of plants may prove effective in producing a diverse array of bioactive diterpenoid natural products. A rice diterpenoid gene cluster is reported that controls the biosynthesis of 5,10-diketo-casbene, a phytoalexin that confers resistance to rice bacterial blight and blast fungus. It recently evolved in the japonica subspecies under artificial selection.

Although natural variations in rice flavonoids exist, and biochemical characterization of a few flavonoid glycosyltransferases has been reported, few studies focused on natural variations in tricin-lignan-glycosides and their underlying genetic basis. In this study, we carried out metabolic profiling of tricin-lignan-glycosides and identified a major quantitative gene annotated as a UDP-dependent glycosyltransferase OsUGT706C2 by metabolite-based genome-wide association analysis. The putative flavonoid glycosyltransferase OsUGT706C2 was characterized as a flavonoid 7-O-glycosyltransferas in vitro and in vivo. Although the in vitro enzyme activity of OsUGT706C2 was similar to that of OsUGT706D1, the expression pattern and induced expression profile of OsUGT706C2 were very different from those of OsUGT706D1. Besides, OsUGT706C2 was specifically induced by UV-B. Constitutive expression of OsUGT706C2 in rice may modulate phenylpropanoid metabolism at both the transcript and metabolite levels. Furthermore, overexpressing OsUGT706C2 can enhance UV-B tolerance by promoting ROS scavenging in rice. Our findings might make it possible to use the glycosyltransferase OsUGT706C2 for crop improvement with respect to UV-B adaptation and/or flavonoid accumulation, which may contribute to stable yield.

Chlorophyll plays remarkable and critical roles in photosynthetic light-harvesting, energy transduction and plant development. In this study, we identified a rice Chl-deficient mutant, ygdl-1 (yellow green and droopy leaf-1), which showed yellow-green leaves throughout plant development with decreased content of Chls and carotene and an increased Chl a/b ratio. The ygdl-1 mutant also exhibited severe defects in chloroplast development, including disorganized grana stacks. Sequence analysis revealed that the mutant contained a T-DNA insertion within the promoter of a fructose-1,6-bisphosphate aldolase (OsAld-Y), which dramatically reduced the OsAld-Y mRNA level, and its identity was verified by transgenic complementation. Real-time PCR analysis showed that the expression levels of genes associated with chlorophyll biosynthesis and chloroplast development were concurrently altered in the ygdl-1 mutant. The expression of OsAld-Y-GFP fusion protein in tobacco epidermal cells showed that OsAld-Y was localized to the peroxisome. In addition, the analysis of primary carbon metabolites revealed the significantly reduced levels of sucrose and fructose in the mutant leaves, while the glucose content was similar to wild-type plants. Our results suggest that the OsAld-Y participates in Chl accumulation, chloroplast development and plant growth by influencing the photosynthetic rate of leaves and the sugar metabolism of rice.

The plant metabolome is characterized by extensive diversity and is often regarded as a bridge between genome and phenome. Here we report metabolic and phenotypic genome-wide studies (mGWAS and pGWAS) in rice grain that, in addition to previous metabolic GWAS in rice leaf and maize kernel, show both distinct and overlapping aspects of genetic control of metabolism within and between species. We identify new candidate genes potentially influencing important metabolic and/or morphological traits. We show that the differential genetic architecture of rice metabolism between different tissues is in part determined by tissue specific expression. Using parallel mGWAS and pGWAS we identify new candidate genes potentially responsible for variation in traits such as grain colour and size, and provide evidence of metabotype-phenotype linkage. Our study demonstrates a powerful strategy for interactive functional genomics and metabolomics in plants, especially the cloning of minor QTLs for complex phenotypic traits., The plant metabolome is often considered as a bridge between genomic and phenotypic variation. Here, the authors perform metabolic and phenotypic genome-wide association studies in rice grain, and compare to prior studies on rice leaf and maize kernel, to demonstrate a new strategy to investigate complex traits.

Highlight The rice putative methyltransferase gene OsTSD2 is functionally identified and is shown to control root development in a way that involves pectin synthesis and modification in a zone-dependent manner., Pectin synthesis and modification are vital for plant development, although the underlying mechanisms are still not well understood. Here, we report the functional characterization of the OsTSD2 gene, which encodes a putative methyltransferase in rice. All three independent T-DNA insertion lines of OsTSD2 displayed dwarf phenotypes and serial alterations in different zones of the root. These alterations included abnormal cellular adhesion and schizogenous aerenchyma formation in the meristematic zone, inhibited root elongation in the elongation zone, and higher lateral root density in the mature zone. Immunofluorescence (with LM19) and Ruthenium Red staining of the roots showed that unesterified homogalacturonan (HG) was increased in Ostsd2 mutants. Biochemical analysis of cell wall pectin polysaccharides revealed that both the monosaccharide composition and the uronic acid content were decreased in Ostsd2 mutants. Increased endogenous ABA content and opposite roles performed by ABA and IAA in regulating cellular adhesion in the Ostsd2 mutants suggested that OsTSD2 is required for root development in rice through a pathway involving pectin synthesis/modification. A hypothesis to explain the relationship among OsTSD2, pectin methylesterification, and root development is proposed, based on pectin’s function in regional cell extension/division in a zone-dependent manner.

The maternal-to-zygotic transition (MZT) is characterized by the turnover of zygote development from maternal to zygotic control, and has been extensively studied in animals. A majority of studies have suggested that early embryogenesis is maternally controlled and that the zygotic genome remains transcriptionally inactive prior to the MZT. However, little is known about the MZT in higher plants, and its timing and impact remain uncharacterized. Here, we constructed cDNA libraries from tobacco (Nicotiana tabacum) egg cells, zygotes and two-celled embryos for gene expression profiling analysis, followed by RT-PCR confirmation. These analyses, together with experiments using zygote microculture coupled with transcription inhibition, revealed that a marked change in transcript profiles occurs approximately 50 h after fertilization, and that the MZT is initiated prior to zygotic division in tobacco. Although maternal transcripts deposited in egg cells support several early developmental processes, they appear to be insufficient for zygotic polar growth and subsequent cell divisions. Thus, we propose that de novo transcripts are probably required to trigger embryogenesis in later zygotes in tobacco.