Metabolomic and Transcriptomic Analyses Reveal the Molecular Mechanism Underlying the Massive Accumulation of Secondary Metabolites in Fenugreek (Trigonella foenum-graecum L.) Seeds.

Fenugreek (Trigonella foenum-graecum L.) is a traditional medicinal plant for treating human diseases that is widely cultivated in many countries. However, the component and related metabolic pathways are still unclear. To understand the changes in expression of the component and related genes during seed development, this study employed metabolomic and transcriptomic analyses and integrative analysis to explore the metabolites and pathways involved in the growth of fenugreek. The antifungal activity of the fenugreek seeds was also analyzed. A total of 9499 metabolites were identified in the positive ion mode, and 8043 metabolites were identified in the negative ion mode. Among them, the main components were fatty acyls, prenol lipids, steroids, steroid derivatives, flavonoids, and isoflavonoids. Among these enriched pathways, the top 20 pathways were "flavone and flavonol biosynthesis", "isoflavonoid biosynthesis", and "flavonoid biosynthesis". 3,7-Di-O-methylquercetin, flavonoids, pseudobaptigenin, isoflavonoids, methylecgonine, alkaloids, and derivatives were the most significantly upregulated metabolites. There were 38,137 differentially expressed genes (DEGs) identified via transcriptomic analysis. According to the KEGG pathway enrichment analysis, 147 DEGs were significantly enriched in "flavonoid biosynthesis". Ten DEGs of the six key enzymes were found to be involved in three pathways related to flavonoid and alkaloid synthesis in fenugreek. The antifungal activity test revealed the inhibitory effect of the ethanol extract of fenugreek seeds on Alternaria tenuissima (Kunze)Wiltshire and Magnaporthe oryzae. These findings further prove that the use of botanical pesticides in fenugreek fruit has research value.

Variation of gene expression of fatty acid acyl CoA reductase associated with wax secretion of a scale insect, Ericerus pela, and identification of its regulation factors through the accessible chromatin analyses and yeast one-hybrid.

The Chinese white wax scale insect (CWWSI), Ericerus pela, can secret an amount of wax equivalent to their body weight. Previous studies demonstrated the fatty acyl-CoA reductase (far3) plays a pivotal role in wax secretion of CWWSI. The high expression of far3 is crucial for the massive wax secretion. However, the transcription regulation of far3 was not clear. To identify regulatory factors that control the expression of far3, the assay for transposase-accessible chromatin (ATAC) and yeast one-hybrid (Y1H) were carried out in this study. The ATAC sequencing of the CWWSI at the early wax-secretion stage ATAC-seq resulted in 22.75 GB raw data, generated 75,827,225 clean reads and revealed 142,771 peaks. There was one significant peak in the 3 kb upstream regulation regions. The peak sequence is located between -1000 and -670 bp upstream of the far3 transcription start site, spanning a length of 331 bp. This peak sequence served as bait for creating the pAbAi-peak recombinant vector, used in Y1H screenings to identify proteins interacting with far3 gene. The results indicate a successful CWWSI cDNA library construction with a capacity of 1.2 × 107 colony forming unit, a 95.8% recombination rate, and insert sizes between 1,000 and 2,000 bp. Self-activation tests established that 100 ng/mL of AbA effectively inhibited bait vector self-activation. Finally, a total of 88 positive clones were selected. After sequencing and removal of duplication, 63 unique clones were obtained from these screened colonies. By aligning the clone sequences with full-length transcriptome and genome of CWWSI, the full-length coding sequences of these clones were obtained. BlastX analysis identified a transcription factor, nuclear transcription factor Y beta, and two co-activators, cAMP-response-element-binding-protein-binding protein and WW domain binding protein 2. Reverse transcription quantitative polymerase chain reaction analysis confirmed that their expression patterns were consistent with the developmental stages preceding wax secretion and matched the wax secretion characteristics during ovulation periods. These results are beneficial for further research into the regulatory mechanisms of wax secretion of CWWSI.

EDIII-Fc induces protective immune responses against the Zika virus in mice and rhesus macaque.

Zika virus can infect the fetus through the placental barrier, causing ZIKV congenital syndrome and even miscarriage, which can cause great harm to pregnant women and infants. Currently, there is no vaccine and drug available to combat the Zika virus. In this study, we designed a fusion protein named EDIII-Fc, including the EDIII region of Zika E protein and human IgG Fc fragment, and obtained 293T cells that stably secreted EDIII-Fc protein using the lentiviral expression system. Mice were immunized with the EDIII-Fc protein, and it was observed that viral replication was significantly inhibited in the immunized mice compared to non-immunized mice. In rhesus macaques, we found that EDIII-Fc effectively induce the secretion of neutralizing antibodies and T cell immunity. These experimental data provide valid data for further use of Zika virus E protein to prepare an effective, safe, affordable Zika vaccine.