Salicylic acid (SA) is a ubiquitous plant hormone with a long history in human civilization^1,2. Because of the central role of SA in orchestrating plant pathogen defence, understanding SA biosynthesis is fundamental to plant immunity research and crop improvement. Isochorismate-derived SA biosynthesis has been well defined in Arabidopsis. However, increasing evidence suggests a crucial function for phenylalanine-derived SA biosynthesis in many other plant species¹. Here we reveal the phenylalanine-derived SA biosynthetic pathway in rice by identifying three dedicated enzymes - peroxisomal benzoyl-CoA:benzyl alcohol benzoyltransferase (BEBT), the endoplasmic reticulum-associated cytochrome P450 enzyme benzylbenzoate hydroxylase (BBH), and cytosolic benzylsalicylate esterase (BSE) that sequentially convert benzoyl-CoA to benzylbenzoate, benzylsalicylate and SA. The pathogen-induced gene expression pattern and SA biosynthetic functions of this triple-enzyme module are conserved in diverse plants. This work fills a major knowledge gap in the biosynthesis of a key plant defence hormone, establishing a foundation for new strategies to create disease-resistant crops.

Salicylic acid (SA) is a prominent defense hormone whose basal level, organ-specific accumulation, and physiological role vary widely among plant species. Of the 2 known pathways of plant SA biosynthesis, the phenylalanine ammonia lyase (PAL) pathway is more ancient and universal but its biosynthetic and physiological roles in diverse plant species remain unclear. Studies in which the PAL pathway is specifically or completely inhibited, as well as a direct comparison of diverse species and different organs within the same species, are needed. To this end, we analyzed the PAL pathway in rice (Oryza sativa) and Arabidopsis (Arabidopsis thaliana), 2 distantly related model plants whose basal SA levels and distributions differ tremendously at the organism and tissue levels. Based on our recent identification of the rice peroxisomal cinnamate:CoA ligases (CNLs), we identified 2 peroxisomal CNLs from Arabidopsis and showed CNL as the most functionally specific enzyme among the known enzymes of the PAL pathway. We then revealed the species- and organ-specific contribution of the PAL pathway to benzoic and salicylic acid biosynthesis and clarified its physiological importance in rice and Arabidopsis. Our findings highlight the necessity to consider species and organ types in future SA-related studies and may help to breed new disease-resistant crops.

Carotenoids are not only photosynthetic and photoprotective pigments in plants, but also essential antioxidative nutrients for human health. The fruit is the main plant organ that synthesizes and sequestrates carotenoids. Fruit ripening is a complicated developmental process, during which the rewiring of the metabolic network is tightly coordinated with the re-organization of cellular and organellular structures. Chili pepper (Capsicum annuum) is one of the major crops that accumulates a distinct level of carotenoids, especially capsanthin, in their ripened fruits. To elucidate how different metabolic and developmental scenarios are regulated in ripening chili pepper fruits, we analyzed the carotenoid profiles and transcriptomes of fruits at different ripening stages. Our pigment analysis indicated an opposite correlation between the contents of carotenoid species with β,β-structures (e.g., β-carotene, zeaxanthin, and capsanthin) and of lutein with the β,ε-structure, whereas lutein displayed a high correlation with chlorophylls during ripening. From the chili pepper Zunla-1 genome, a full repertoire of 38 homologous genes encoding enzymes in the carotenoid biosynthetic pathway was identified. The fluctuations in their transcript abundances during ripening suggested different involvement of these genes in the regulation of carotenoid biosynthesis. We further searched genes of which the expression showed high correlations with the accumulation of β-carotene during the ripening process. Moreover, from the transcriptomic analysis, a total of 17 transcription factors that co-expressed with different groups of carotenoid biosynthetic genes were identified.

Protein structure serves as a critical bridge between sequence and functional annotation, particularly in establishing functional links among distantly homologous proteins with low sequence similarities. However, systematic protein structure-based functional annotations have been lacking in plants, where functions for a significant portion of the proteomes are still elusive. In this study, we leveraged protein structural data from 17 angiosperms to uncover previously unannotated protein functions in plants. After structural clustering, we used the plant clusters to query the UniProtKB/Swiss-Prot database (the expertly curated component of UniProtKB), a repository of expertly curated and reliably annotated proteins, and identified structural matches for thousands of plant clusters that were undetectable by sequence-based BLAST searches. We further selected 120 clusters, which are highly reliable in structural quality and alignment and are well-conserved across plant species, and uncovered various protein functions that are potentially widely important in plants. Finally, we experimentally analyzed one plant cluster structurally resembling the yeast peroxisomal peroxin 8 (PEX8) protein and verified that plant PEX8-like proteins can functionally complement yeast pex8 mutants. Our findings highlight the power of structural comparison in uncovering protein functions in plants.