Chromosome-level genome assembly and functional characterization of terpene synthases provide insights into the volatile terpenoid biosynthesis of Wurfbainia villosa.

Wurfbainia villosa is a well-known medicinal and edible plant that is widely cultivated in the Lingnan region of China. Its dried fruits (called Fructus Amomi) are broadly used in traditional Chinese medicine for curing gastrointestinal diseases and are rich in volatile terpenoids. Here, we report a high-quality chromosome-level genome assembly of W. villosa with a total size of approximately 2.80 Gb, 42 588 protein-coding genes, and a very high percentage of repetitive sequences (87.23%). Genome analysis showed that W. villosa likely experienced a recent whole-genome duplication event prior to the W. villosa-Zingiber officinale divergence (approximately 11 million years ago), and a recent burst of long terminal repeat insertions afterward. The W. villosa genome enabled the identification of 17 genes involved in the terpenoid skeleton biosynthesis pathway and 66 terpene synthase (TPS) genes. We found that tandem duplication events have an important contribution to the expansion of WvTPSs, which likely drove the production of volatile terpenoids. In addition, functional characterization of 18 WvTPSs, focusing on the TPS-a and TPS-b subfamilies, showed that most of these WvTPSs are multi-product TPS and are predominantly expressed in seeds. The present study provides insights into the genome evolution and the molecular basis of the volatile terpenoids diversity in W. villosa. The genome sequence also represents valuable resources for the functional gene research and molecular breeding of W. villosa.

A Comparison of Two Monoterpenoid Synthases Reveals Molecular Mechanisms Associated With the Difference of Bioactive Monoterpenoids Between Amomum villosum and Amomum longiligulare.

The fruits of Amomum villosum and Amomum longiligulare are both used medicinally as Fructus Amomi the famous traditional Chinese medicine, however, the medicinal quality of A. villosum is better than that of A. longiligulare. Volatile terpenoids in the seeds, especially bornyl acetate and borneol, are the medicinal components of Fructus Amomi. The volatile terpenoids and transcriptome of developing seeds of A. villosum and A. longiligulare were compared in this study. The result revealed that the bornyl acetate and borneol contents were higher in A. villosum than in A. longiligulare. Additionally, six terpenoid synthase genes (AlTPS1-AlTPS6) were screened from the transcriptome of A. longiligulare, and AlTPS2 and AlTPS3 were found to share 98 and 83% identity with AvTPS2 and AvBPPS (bornyl diphosphate synthase) from A. villosum, respectively. BPPS is the key enzyme for the biosynthesis of borneol and bornyl acetate. Biochemical assays improved that AlTPS2 had an identical function to AvTPS2 as linalool synthase; however, AlTPS3 produced camphene as the major product and bornyl diphosphate (BPP) as the secondary product, whereas AvBPPS produced BPP as its major product. There was only one different amino acid between AlTPS3 (A496) and AvBPPS (G495) in their conserved motifs, and the site-directed mutation of A496G in DTE motif of AlTPS3 changed the major product from camphene to BPP. Molecular docking suggests that A496G mutation narrows the camphene-binding pocket and decreases the BPP-binding energy, thus increases the product BPP selectivity of enzyme. In addition, the expression level of AvBPPS was significantly higher than that of AlTPS3 in seeds, which was consistent with the related-metabolites contents. This study provides insight into the TPS-related molecular bases for the biosynthesis and accumulation differences of the bioactive terpenoids between A. villosum and A. longiligulare. BPPS, the key gene involved in the biosynthesis of the active compound, was identified as a target gene that could be applied for the quality-related identification and breeding of Fructus Amomi.

Ginsenoside Rd inhibits IL-1ß-induced inflammation and degradation of intervertebral disc chondrocytes by increasing IL1RAP ubiquitination.

Many compounds of ginsenosides show anti-inflammatory properties. However, their anti-inflammatory effects in intervertebral chondrocytes in the presence of inflammatory factors have never been shown. Increased levels of pro-inflammatory cytokines are generally associated with the degradation and death of chondrocytes; therefore, finding an effective and nontoxic substance that attenuates the inflammation is worthwhile. In this study, chondrocytes were isolated from the nucleus pulposus tissues, and the cells were treated with ginsenoside compounds and IL-1ß, alone and in combination. Cell viability and death rate were assessed by CCK-8 and flow cytometry methods, respectively. PCR, western blot, and immunoprecipitation assays were performed to determine the mRNA and protein expression, and the interactions between proteins, respectively. Monomeric component of ginsenoside Rd had no toxicity at the tested range of concentrations. Furthermore, Rd suppressed the inflammatory response of chondrocytes to interleukin (IL)-1ß by suppressing the increase in IL-1ß, tumor necrosis factor (TNF)-α, IL-6, COX-2, and inducible nitric oxide synthase (iNOS) expression, and retarding IL-1ß-induced degradation of chondrocytes by improving cell proliferation characteristics and expression of aggrecan and COL2A1. These protective effects of Rd were associated with ubiquitination of IL-1 receptor accessory protein (IL1RAP), blocking the stimulation of IL-1ß to NF-κB. Bioinformatics analysis showed that NEDD4, CBL, CBLB, CBLC, and ITCH most likely target IL1RAP. Rd increased intracellular ITCH level and the amount of ITCH attaching to IL1RAP. Thus, IL1RAP ubiquitination promoted by Rd is likely to occur by up-regulation of ITCH. In summary, Rd inhibited IL-1ß-induced inflammation and degradation of intervertebral disc chondrocytes by increasing IL1RAP ubiquitination.

Ginsenoside Rd inhibits IL-1ß-induced inflammation and degradation of intervertebral disc chondrocytes by increasing IL1RAP ubiquitination

Many compounds of ginsenosides show anti-inflammatory properties. However, their anti-inflammatory effects in intervertebral chondrocytes in the presence of inflammatory factors have never been shown. Increased levels of pro-inflammatory cytokines are generally associated with the degradation and death of chondrocytes; therefore, finding an effective and nontoxic substance that attenuates the inflammation is worthwhile. In this study, chondrocytes were isolated from the nucleus pulposus tissues, and the cells were treated with ginsenoside compounds and IL-1β, alone and in combination. Cell viability and death rate were assessed by CCK-8 and flow cytometry methods, respectively. PCR, western blot, and immunoprecipitation assays were performed to determine the mRNA and protein expression, and the interactions between proteins, respectively. Monomeric component of ginsenoside Rd had no toxicity at the tested range of concentrations. Furthermore, Rd suppressed the inflammatory response of chondrocytes to interleukin (IL)-1β by suppressing the increase in IL-1β, tumor necrosis factor (TNF)-α, IL-6, COX-2, and inducible nitric oxide synthase (iNOS) expression, and retarding IL-1β-induced degradation of chondrocytes by improving cell proliferation characteristics and expression of aggrecan and COL2A1. These protective effects of Rd were associated with ubiquitination of IL-1 receptor accessory protein (IL1RAP), blocking the stimulation of IL-1β to NF-κB. Bioinformatics analysis showed that NEDD4, CBL, CBLB, CBLC, and ITCH most likely target IL1RAP. Rd increased intracellular ITCH level and the amount of ITCH attaching to IL1RAP. Thus, IL1RAP ubiquitination promoted by Rd is likely to occur by up-regulation of ITCH. In summary, Rd inhibited IL-1β-induced inflammation and degradation of intervertebral disc chondrocytes by increasing IL1RAP ubiquitination.

Meta-analysis of randomized controlled trials for the efficacy and safety of anti-interleukin-13 therapy with lebrikizumab in patients with uncontrolled asthma.

BACKGROUND: Several studies have evaluated the efficacy and safety of lebrikizumab treatment with uncontrolled asthma. However, most of these studies were small and conclusions were inconsistent. Furthermore, whether serum periostin can act as a good predictor of the response to lebrikizumab treatment is still not certain. METHOD: We conducted a systematic review and meta-analysis of randomized controlled trials to evaluate the efficacy and safety of lebrikizumab treatment with uncontrolled asthma. Trials were searched in PubMed, Embase, Web of Science and Cochrane. Outcome measures were the rate of asthma exacerbations, relative changes in the forced expiratory volume in the first second of expiration (FEV1) of predicted value (%) and incidence of adverse events. RESULT: Five trials were finally included. Compared with placebo lebrikizumab treatment significantly decreased the rate of exacerbations(risk ratio [RR] 0.66 [95% confidence interval {CI}, 0.54-0.80]; p < 0.0001; n = 2039) and increased FEV1% of predicted value (weighted mean difference [WMD] 5.46 [95% CI, 2.48-8.43]; p < 0.0003; n = 351). Patients with high levels of serum periostin had greater exacerbation rate reductions (RR 0.59 [95%CI, 0.50-0.70]; p < 0.00001; n = 1157) and FEV1 of predicted value improvement (WMD 7.18 [95% CI, 2.93-11.42]; p < 0.0009; n = 177) than patients with low periostin levels in exacerbation rate reductions (RR 0.73 [95% CI, 0.47-1.14]; p < 0.17; n = 882) and FEV1 of predicted value improvement (WMD3.79 [95% CI, 0.39-7.97]; p < 0.08; n = 174). There was no significant difference in the incidence of adverse events in patients with lebrikizumab compared to placebo (RR 1.03 [95% CI, 0.99-1.06]; p < 0.11; n = 2056). CONCLUSION: In patients with uncontrolled asthma, lebrikizumab treatment significantly decreased the rate of exacerbation and improved lung function, especially for patients with high periostin levels.

Characterization and functional analysis of the genes encoding 1-deoxy-D-xylulose-5-phosphate reductoisomerase and 1-deoxy-D-xylulose-5-phosphate synthase, the two enzymes in the MEP pathway, from Amomum villosum Lour.

A DXR gene, AvDXR (GenBank accession no. FJ459894), and a DXS gene, AvDXS (GenBank accession no. FJ455512), were isolated from the leaves of Amomum villosum, one of the most well-known and authentic herbs in South China. The 1,749-bp full-length cDNA of AvDXR encoded a peptide of 472 amino acids, and the 2,347-bp full-length cDNA of AvDXS encoded a peptide of 715 amino acids. The deduced amino acid sequences of the AvDXR and AvDXS proteins share high homology with DXRs and DXSs from other plant species, and AvDXS belongs to class 1 plant DXS. The characterization based on bioinformatic analysis indicated that the AvDXR and AvDXS encoded functional proteins as DXR and DXS, respectively. The functional color assay in Escherichia coli with pAC-BETA implied that AvDXR and AvDXS encoded functional proteins that manipulated the biosynthesis of isoprenoid precursors. Both AvDXR and AvDXS were expressed extensively in the leaves, stems, roots, pericarps and seeds of A. villosum. AvDXS expression was similar in all tissues investigated, whereas higher levels of AvDXR were observed in the fruits, the main part for the accumulation of volatile oil in this plant. AvDXR was transformed into tobacco to confirm its function further. Overexpression of AvDXR in transgenic T1 generation tobacco increased DXR activity, photosynthetic pigment content and volatile isoprenoid components, and the increase of photosynthetic pigment content was consistent with the AvDXR transcription level. This study demonstrated that AvDXR plays important role in isoprenoid biosynthesis and it is useful for metabolic engineering.