Search Results (8)

Huperzine A (HupA) is a natural Lycopodium alkaloid known for its potent neuroprotective properties through the inhibition of acetylcholinesterase. Nevertheless, the limited understanding of its biosynthesis restricts its broader application. This study integrates full-length and second-generation transcriptomes with the quantification of HupA and its precursor, huperzine B, across various tissues of Huperzia serrata, the primary source plant. By employing phylogenetic clustering, expression profiling, and correlation analysis between gene expression and metabolite abundance, we identified 71 candidate genes from seven enzyme families potentially involved in the synthesis of the HupA backbone, including lysine/ornithine decarboxylases, copper amine oxidases (CAOs), chalcone synthases, and cytochrome P450 monooxygenases. Additionally, 28 genes from two families were identified for modification reactions, specifically 2-oxoglutarate/Fe(II)-dependent dioxygenases and caffeoyl shikimate esterases. Comparative analysis between young and mature leaves revealed 3801 genes with higher expression in young leaves, with 84 showing a high correlation with HupA content across seven families. Protein–protein interaction network analysis indicated possible interactions with transcription factors from the MYB, NF-YC, GRAS, ERF, BHLH, and SAP families. Functional validation of two candidate CAOs in planta confirmed their catalytic roles in amine/alkaloid metabolism. This study provides a theoretical foundation and a set of candidate genes for elucidating the biosynthetic pathway of HupA and related alkaloids in H. serrata. Full article

Butylphthalide, a prescription medicine recognized for its efficacy in treating ischemic strokes approved by the State Food and Drug Administration of China in 2005, is sourced from the traditional botanical remedy Ligusticum chuanxiong. While chemical synthesis offers a viable route, limitations in the production of isomeric variants with compromised bioactivity necessitate alternative strategies. Addressing this issue, biosynthesis offers a promising solution. However, the intricate in vivo pathway for butylphthalide biosynthesis remains elusive. In this study, we examined the distribution of butylphthalide across various tissues of L. chuanxiong and found a significant accumulation in the rhizome. By searching transcriptome data from different tissues of L. chuanxiong, we identified four rhizome-specific genes annotated as 2-oxoglutarate-dependent dioxygenase (2-OGDs) that emerged as promising candidates involved in butylphthalide biosynthesis. Among them, LcSAO1 demonstrates the ability to catalyze the desaturation of senkyunolide A at the C-4 and C-5 positions, yielding the production of butylphthalide. Experimental validation through transient expression assays in Nicotiana benthamiana corroborates this transformative enzymatic activity. Notably, phylogenetic analysis of LcSAO1 revealed that it belongs to the DOXB clade, which typically encompasses genes with hydroxylation activity, rather than desaturation. Further structure modelling and site-directed mutagenesis highlighted the critical roles of three amino acid residues, T98, S176, and T178, in substrate binding and enzyme activity. By unraveling the intricacies of the senkyunolide A desaturase, the penultimate step in the butylphthalide biosynthesis cascade, our findings illuminate novel avenues for advancing synthetic biology research in the realm of medicinal natural products. Full article

The 2-oxoglutarate and Fe (II)-dependent dioxygenase (2ODD-C) family of 2-oxoglutarate-dependent dioxygenases potentially participates in the biosynthesis of various metabolites under various abiotic stresses. However, there is scarce information on the expression profiles and roles of 2ODD-C genes in Camellia sinensis. We identified 153 Cs2ODD-C genes from C. sinensis, and they were distributed unevenly on 15 chromosomes. According to the phylogenetic tree topology, these genes were divided into 21 groups distinguished by conserved motifs and an intron/exon structure. Gene-duplication analyses revealed that 75 Cs2ODD-C genes were expanded and retained after WGD/segmental and tandem duplications. The expression profiles of Cs2ODD-C genes were explored under methyl jasmonate (MeJA), polyethylene glycol (PEG), and salt (NaCl) stress treatments. The expression analysis showed that 14, 13, and 49 Cs2ODD-C genes displayed the same expression pattern under MeJA and PEG treatments, MeJA and NaCl treatments, and PEG and NaCl treatments, respectively. A further analysis showed that two genes, Cs2ODD-C36 and Cs2ODD-C21, were significantly upregulated and downregulated after MeJA, PEG, and NaCl treatments, indicating that these two genes played positive and negative roles in enhancing the multi-stress tolerance. These results provide candidate genes for the use of genetic engineering technology to modify plants by enhancing multi-stress tolerance to promote phytoremediation efficiency. Full article

The 2-oxoglutarate and Fe(II)-dependent dioxygenase (2OGD) superfamily is the second largest enzyme family in the plant genome, and its members are involved in various oxygenation and hydroxylation reactions. Due to their important biochemical significance in metabolism, a systematic analysis of the plant 2OGD genes family is necessary. Here, we identified 160, 179, and 337 putative 2OGDs from Brassica rapa, Brassica oleracea, and Brassica napus. According to their gene structure, domain, phylogenetic features, function, and previous studies, we also divided 676 2OGDs into three subfamilies: DOXA, DOXB, and DOXC. Additionally, homologous and phylogenetic comparisons of three subfamily genes provided valuable insight into the evolutionary characteristics of the 2OGD genes from Brassica plants. Expression profiles derived from the transcriptome and Genevestigator database exhibited distinct expression patterns of the At2OGD, Br2OGD, and Bo2OGD genes in different developmental stages, tissues, or anatomical parts. Some 2OGD genes showed high expression levels in various tissues, such as callus, seed, silique, and root tissues, while other 2OGD genes were expressed at very low levels in other tissues. Analysis of six Bo2OGD genes in different tissues by qRT-PCR indicated that these genes are involved in the metabolism of gibberellin, which in turn regulates plant growth and development. Our working system analysed 2OGD gene families of three Brassica plants and laid the foundation for further study of their functional characterization. Full article

The FTO protein is involved in a wide range of physiological processes, including adipogenesis and osteogenesis. This two-domain protein belongs to the AlkB family of 2-oxoglutarate (2-OG)- and Fe(II)-dependent dioxygenases, displaying N⁶-methyladenosine (N⁶-meA) demethylase activity. The aim of the study was to characterize the relationships between the structure and activity of FTO. The effect of cofactors (Fe²⁺/Mn²⁺ and 2-OG), Ca²⁺ that do not bind at the catalytic site, and protein concentration on FTO properties expressed in either E. coli (^ECFTO) or baculovirus (^BESFTO) system were determined using biophysical methods (DSF, MST, SAXS) and biochemical techniques (size-exclusion chromatography, enzymatic assay). We found that ^BESFTO carries three phosphoserines (S184, S256, S260), while there were no such modifications in ^ECFTO. The S256D mutation mimicking the S256 phosphorylation moderately decreased FTO catalytic activity. In the presence of Ca²⁺, a slight stabilization of the FTO structure was observed, accompanied by a decrease in catalytic activity. Size exclusion chromatography and MST data confirmed the ability of FTO from both expression systems to form homodimers. The MST-determined dissociation constant of the FTO homodimer was consistent with their in vivo formation in human cells. Finally, a low-resolution structure of the FTO homodimer was built based on SAXS data. Full article

Gibberellins (GAs), a class of diterpenoid phytohormones, play a key role in regulating diverse processes throughout the life cycle of plants. Bioactive GA levels are rapidly regulated by Gibberellin-dioxygenases (GAox), which are involved in the biosynthesis and deactivation of gibberellin. In this manuscript, a comprehensive genome-wide analysis was carried out to find all GAox in Camellia sinensis. For the first time in a tea plant, 14 CsGAox genes, containing two domains, DIOX_N (PF14226) and 2OG-FeII_Oxy, were identified (PF03171). These genes all belong to 2-oxoglutarate-dependent dioxygenases (2-ODD), including four CsGA20ox (EC: 1.14.11.12), three CsGA3ox (EC: 1.14.11.15), and seven CsGA2ox (EC: 1.14.11.13). According to the phylogenetic classification as in Arabidopsis, the CsGAox genes spanned five subgroups. Each CsGAox shows tissue-specific expression patterns, although these vary greatly. Some candidate genes, which may play an important role in response to external abiotic stresses, have been identified with regards to patterns, such as CsGA20ox2, CsGA3ox2, CsGA3ox3, CsGA2ox1, CsGA2ox2, and CsGA2ox4. The bioactive GA levels may be closely related to the GA20ox, GA3ox and GA2ox genes. In addition, the candidate genes could be used as marker genes for abiotic stress resistance breeding in tea plants. Full article

Flavonoids are secondary metabolites derived from phenylalanine and acetate metabolism. They fulfil a variety of functions in plants and have health benefits for humans. During the synthesis of the tricyclic flavonoid natural products in plants, oxidative modifications to the central C ring are catalyzed by four of Fe^II and 2-oxoglutarate dependent (2-ODD) oxygenases, namely flavone synthase I (FNS I), flavonol synthase (FLS), anthocyanidin synthase (ANS) and flavanone 3β-hydroxylase (FHT). FNS I, FLS and ANS are involved in desaturation of C2–C3 of flavonoids and FHT in hydroxylation of C3. FNS I, which is restricted to the Apiaceae species and in rice, is predicted to have evolved from FHT by duplication. Due to their sequence similarity and substrate specificity, FLS and ANS, which interact with the α surface of the substrate, belong to a group of dioxygenases having a broad substrate specificity, while FNS I and FHT are more selective, and interact with the naringenin β surface. Here, we summarize recent findings regarding the function of the four 2-ODD oxygenases and the relationship between their catalytic activity, their polypeptide sequence and their tertiary structure. Full article

Flavonoids and biochemically-related chalcones are important secondary metabolites, which are ubiquitously present in plants and therefore also in human food. They fulfill a broad range of physiological functions in planta and there are numerous reports about their physiological relevance for humans. Flavonoids have in common a basic C₆-C₃-C₆ skeleton structure consisting of two aromatic rings (A and B) and a heterocyclic ring (C) containing one oxygen atom, whereas chalcones, as the intermediates in the formation of flavonoids, have not yet established the heterocyclic C-ring. Flavonoids are grouped into eight different classes, according to the oxidative status of the C-ring. The large number of divergent chalcones and flavonoid structures is from the extensive modification of the basic molecules. The hydroxylation pattern influences physiological properties such as light absorption and antioxidative activity, which is the base for many beneficial health effects of flavonoids. In some cases antiinfective properties are also effected. Full article