Background Karst-adapted plant, Lysionotus pauciflours accumulates special secondary metabolites with a wide range of pharmacological effects for surviving in drought and high salty areas, while researchers focused more on their environmental adaptations and evolutions. Nevadensin (5,7-dihydroxy-6,8,4'-trimethoxyflavone), the main active component in L. pauciflours, has unique bioactivity of such as anti-inflammatory, anti-tubercular, and anti-tumor or cancer. Complex decoration of nevadensin, such as hydroxylation and glycosylation of the flavone skeleton determines its diversity and biological activities. The lack of omics data limits the exploration of accumulation mode and biosynthetic pathway. Herein, we integrated transcriptomics, metabolomics, and microbial recombinant protein system to reveal hydroxylation and glycosylation involving nevadensin biosynthesis in L. pauciflours. Results Up to 275 flavonoids were found to exist in L. pauciflorus by UPLC-MS/MS based on widely targeted metabolome analysis. The special flavone nevadensin (5,7-dihydroxy-6,8,4'-trimethoxyflavone) is enriched in different tissues, as are its related glycosides. The flavonoid biosynthesis pathway was drawn based on differential transcripts analysis, including 9 PAL, 5 C4H, 8 4CL, 6 CHS, 3 CHI, 1 FNSII, and over 20 OMTs. Total 310 LpCYP450s were classified into 9 clans, 36 families, and 35 subfamilies, with 56% being A-type CYP450s by phylogenetic evolutionary analysis. According to the phylogenetic tree with AtUGTs, 187 LpUGTs clustered into 14 evolutionary groups (A-N), with 74% being E, A, D, G, and K groups. Two LpCYP82D members and LpUGT95 were functionally identified in Saccharomyces cerevisiae and Escherichia coli, respectively. CYP82D-8 and CYP82D-1 specially hydroxylate the 6- or 8-position of A ring in vivo and in vitro, dislike the function of F6H or F8H discovered in basil which functioned depending on A-ring substituted methoxy. These results refreshed the starting mode that apigenin can be firstly hydroxylated on A ring in nevadensin biosynthesis. Furthermore, LpUGT95 clustered into the 7-OGT family was verified to catalyze 7-O glucosylation of nevadensin accompanied with weak nevadensin 5-O glucosylation function, firstly revealed glycosylation modification of flavones with completely substituted A-ring. Conclusions Metabolomic and full-length transcriptomic association analysis unveiled the accumulation mode and biosynthetic pathway of the secondary metabolites in the karst-adapted plant L. pauciflorus. Moreover, functional identification of two LpCYP82D members and one LpUGT in microbe reconstructed the pathway of nevadensin biosynthesis.
<i>Periploca forrestii</i>, a medicinal plant of the family Apocynaceae, is known as an effective and widely used clinical prescription for the treatment of rheumatoid diseases. In this study, we de novo sequenced and assembled the completement chloroplast (cp) genome of <i>P. forrestii</i> based on combined Oxford Nanopore PromethION and Illumina data. The cp genome was 153,724 bp in length and had four subregions. Moreover, an 84,433-bp large single-copy (LSC) and a 17,731-bp small single-copy (SSC) were separated by 25,780-bp inverted repeats (IRs). The cp genome included 132 genes with 18 duplicates in the IRs. A total of 45 repeat structures and 183 simple sequence repeats (SSRs) were detected. Codon usage showed a bias toward A/T-ending codons. A comparative study of Apocynaceae revealed that an IR expansion occurred on P. forrestii. The Ka/Ks values of eight species of Apocynaceae suggested that positive selection was exerted on <i>psaI</i> and <i>ycf2</i>, which might reflect specific adaptions to the <i>P. forrestii</i> particular growth environment. Phylogenetic analysis indicated that Periplocoideae was a sister to Asclepiadoideae, forming a monophyletic group in the family Apocynaceae. This study provided an important <i>P. forrestii</i> genomic resource for future evolutionary studies and the phylogenetic reconstruction of the family Apocynaceae.
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