Juglandaceae (walnut) is made up of several economically and ecologically valuable tree species. Chloroplasts, vitally important for plant growth, are also a rich source of genetic and evolutionary information. Both mutational pressure and natural selection are drivers of codon usage pattern variation among genes. Here, we studied the codon usage of Juglandaceae chloroplast genomes in order to further our understanding of the biology and evolution of this plant family. The codon usage patterns associated with the chloroplast genomes of 26 Juglandaceae samples were analyzed. Short CDS sequences (<100 amino acids) and sequencing containing internal stop codons were removed from comparative analyses. The contents of uracil (U) (31.5%–32.0%) and adenine (A) (30.0%–31.2%) of all 26 samples were higher than those of cytosine (C) (17.2%–17.7%) and guanine (G) 19.9%–20.7%. According to the neutrality and correspondence analyses, chloroplast codons tended to exhibit conserved GC content and were primarily altered by natural selection. The parity rule 2 plot analysis revealed that AU were more common than GC at the third-codon position. According to the effective number of codon (ENC) plot analysis, codon preference was driven by natural selection and protein translation, among other factors. This study represents the first examination of the codon usage characteristics of Juglandaceae plants, as revealed through the study of codon bias in 26 Juglandaceae samples.

Camellia oleifera, a widely cultivated woody oil crop, holds economic significance because of its ability to grow without encroaching on cultivated land. The pericarp of C. oleifera is abundant in flavonoids and phenolic acids, which offer significant nutritional benefits. This study used metabolomic technology (UPLC-ESI-MS/MS) to discern metabolite variances in the pericarp of three C. oleifera types (COT, BFOT, and SFOT) during the maturity stage and subsequently analyzed and compared them. A total of 1117 metabolites were detected in the study, including 277 flavonoids, 221 phenolic acids, 108 lipids, 93 amino acids and their derivatives, 83 organic acids, 59 nucleotides and their derivatives, 57 alkaloids, 52 lignans, 44 tannins, 23 terpenoids, and 100 miscellaneous metabolites (such as sugars, alcohols, vitamins, and other unclassified substances). Clustering and PCA analyses revealed distinct separation of COT, BFOT, and SFOT, indicating variances in metabolites within the pericarp peels of these three C. oleifera types. KEGG enrichment analysis demonstrated that 143 shared differential metabolites were primarily associated with amino acid biosynthesis. These findings are expected to significantly enhance the current knowledge of the C. oleifera pericarp and pave the way for future development and use efforts.

Phoebe bournei is a precioustimber species and is listed as a national secondary protection plant in China. However, seedlings show obvious photosynthetic declinewhen grown long-term under an elevated CO₂ concentration (eCO₂). The global CO₂ concentration is predicted to reach 700 μmol·mol⁻¹ by the end of this century; however, little is known about what causes the photosynthetic decline of P. bournei seedlings under eCO₂ or whether this photosynthetic decline could be controlled by fertilization measures. To explore this problem, one-year-old P. bournei seedlings were grown in an open-top air chamber under either an ambient CO₂ (aCO₂) concentration (350 ± 70 μmol·mol⁻¹) or an eCO₂ concentration (700 ± 10 μmol·mol⁻¹) from June 12th to September 8th and cultivated in soil treated with either moderate (0.8 g per seedling) or high applications (1.2 g per seedling) of nitrate or ammonium. Under eCO₂, the net photosynthetic rate (Pn) of P. bournei seedlings treated with a moderate nitrate application was 27.0% lower than that of seedlings grown under an aCO₂ concentration (p < 0.05), and photosynthetic declineappeared to be accompanied by a reduction of the electron transport rate (ETR), actual photochemical efficiency, chlorophyll content, ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco), rubisco activase (RCA) content, leaf thickness, and stomatal density. The Pn of seedlings treated with a high application of nitrate under eCO₂ was 5.0% lower than that of seedlings grown under aCO₂ (p > 0.05), and photosynthetic declineoccurred more slowly, accompanied by a significant increase in rubisco content, RCA content, and stomatal density. The Pn of P. bournei seedlings treated with either a moderate or a high application of ammonium and grown under eCO₂ was not significantly differentto that of seedlings grown under aCO₂—there was no photosynthetic decline—and the ETR, chlorophyll content, rubisco content, RCA content, and leaf thickness values were all increased. Increasing the application of nitrate or the supply of ammonium could slow down or prevent the photosynthetic declineof P. bournei seedlings under eCO₂ by changing the leaf structure and photosynthetic physiological characteristics.

The YABBY gene family is a plant transcription factor that exists in all seed plants. YABBY family members have been studied extensively in various plants and were to play significant roles in plant growth and development. Juglans, especially walnuts, are important economic tree species that are widely distributed worldwide. However, the identification and related research of YABBY in Juglans have not been reported to date. In this study, we identified 19 YABBY genes from two Juglans species, namely, J. regia and J. mandshurica. Ten JrYABBY genes and nine JmYABBY genes were divided into five subfamilies (YAB1/3, YAB2, INO, CRC, and YAB5). Sequence analysis revealed that all encoded YABBY protein sequences had a highly conserved YABBY and C2C2 zinc-finger domains. An analysis of the assumed cis-acting elements revealed that JrYABBY and JmYABBY genes were deeply involved in phytohormone and light responses. Further, gene expression pattern analysis suggested that most walnut YABBY genes were likely involved in peel and flower development and responses to biotic stress. This study not only suppled novel insights into the evolutionary basis of YABBY gene families in Juglans, but also provided clues for the further functional verification and investigation of YABBY genes in other tree species.

Coumarins in the pericarp of Zanthoxylum contribute to the characteristic numbing–aromatic flavor and are associated with diverse bioactivities. To characterize coumarin divergence between two Zanthoxylum materials, mature pericarps of Dahongpao Z. bungeanum (red Sichuan pepper) and Z. planispinum var. dingtanensis (green Sichuan pepper) were analyzed by widely targeted UPLC–ESI–MS/MS metabolomics integrated with transcriptome sequencing. This approach enabled joint profiling of metabolites and transcripts to identify genes associated with material-specific coumarin accumulation. Across the two materials, 583 metabolites were detected, with flavonoids, phenolic acids, and alkaloids as the predominant classes. Among these, 24 coumarins were identified, and most showed significantly higher abundance in green Sichuan pepper than in red Sichuan pepper. Pathway enrichment analysis indicated that differentially accumulated coumarins were mainly associated with the phenylpropanoid biosynthesis pathway, consistent with coordinated metabolic and transcriptional regulation. The integration of metabolite abundance with gene expression patterns identified 56 candidate genes strongly correlated with scopoletin and scopolin accumulation. To evaluate functional relevance, CCoAOMT, COMT, and F6H were cloned and transiently overexpressed in Nicotiana benthamiana. Transient expression assays showed that overexpression of each gene increased scopoletin and scopolin, supporting their involvement in coumarin biosynthesis. Collectively, these results clarify molecular determinants of coumarin variation between the two materials and highlight candidate genes for quality improvement and metabolic engineering.