Identification of key genes involved in secondary metabolite biosynthesis in Digitalis purpurea.

The medicinal plant Digitalis purpurea produces cardiac glycosides that are useful in the pharmaceutical industry. These bioactive compounds are in high demand due to ethnobotany's application to therapeutic procedures. Recent studies have investigated the role of integrative analysis of multi-omics data in understanding cellular metabolic status through systems metabolic engineering approach, as well as its application to genetically engineering metabolic pathways. In spite of numerous omics experiments, most molecular mechanisms involved in metabolic pathways biosynthesis in D. purpurea remain unclear. Using R Package Weighted Gene Co-expression Network Analysis, co-expression analysis was performed on the transcriptome and metabolome data. As a result of our study, we identified transcription factors, transcriptional regulators, protein kinases, transporters, non-coding RNAs, and hub genes that are involved in the production of secondary metabolites. Since jasmonates are involved in the biosynthesis of cardiac glycosides, the candidate genes for Scarecrow-Like Protein 14 (SCL14), Delta24-sterol reductase (DWF1), HYDRA1 (HYD1), and Jasmonate-ZIM domain3 (JAZ3) were validated under methyl jasmonate treatment (MeJA, 100 µM). Despite early induction of JAZ3, which affected downstream genes, it was dramatically suppressed after 48 hours. SCL14, which targets DWF1, and HYD1, which induces cholesterol and cardiac glycoside biosynthesis, were both promoted. The correlation between key genes and main metabolites and validation of expression patterns provide a unique insight into the biosynthesis mechanisms of cardiac glycosides in D. purpurea.

System network analysis of Rosmarinus officinalis transcriptome and metabolome-Key genes in biosynthesis of secondary metabolites.

Medicinal plants contain valuable compounds that have attracted worldwide interest for their use in the production of natural drugs. The presence of compounds such as rosmarinic acid, carnosic acid, and carnosol in Rosmarinus officinalis has made it a plant with unique therapeutic effects. The identification and regulation of the biosynthetic pathways and genes will enable the large-scale production of these compounds. Hence, we studied the correlation between the genes involved in biosynthesis of the secondary metabolites in R. officinalis using proteomics and metabolomics data by WGCNA. We identified three modules as having the highest potential for the metabolite engineering. Moreover, the hub genes highly connected to particular modules, TFs, PKs, and transporters were identified. The TFs of MYB, C3H, HB, and C2H2 were the most likely candidates associated with the target metabolic pathways. The results indicated that the hub genes including Copalyl diphosphate synthase (CDS), Phenylalanine ammonia lyase (PAL), Cineole synthase (CIN), Rosmarinic acid synthase (RAS), Tyrosine aminotransferase (TAT), Cinnamate 4-hydroxylase (C4H), and MYB58 are responsible for biosynthesis of important secondary metabolites. Thus, we confirmed these results using qRT-PCR after treating R. officinalis seedlings with methyl jasmonate. These candidate genes may be employed for genetic and metabolic engineering research to increase R. officinalis metabolite production.

Integration of transcriptomic and metabolomic analyses provides insights into response mechanisms to nitrogen and phosphorus deficiencies in soybean.

Nitrogen (N) and phosphorus (P) are two essential plant macronutrients that can limit plant growth by different mechanisms. We aimed to shed light on how soybean respond to low nitrogen (LN), low phosphorus (LP) and their combined deficiency (LNP). Generally, these conditions triggered changes in gene expression of the same processes, including cell wall organization, defense response, response to oxidative stress, and photosynthesis, however, response was different in each condition. A typical primary response to LN and LP was detected also in soybean, i.e., the enhanced uptake of N and P, respectively, by upregulation of genes for the corresponding transporters. The regulation of genes involved in cell wall organization showed that in LP roots tended to produce more casparian strip, in LN more secondary wall biosynthesis occurred, and in LNP reduction in expression of genes involved in secondary wall production accompanied by cell wall loosening was observed. Flavonoid biosynthesis also showed distinct pattern of regulation in different conditions: more anthocyanin production in LP, and more isoflavonoid production in LN and LNP, which we confirmed also on the metabolite level. Interestingly, in soybean the nutrient deficiencies reduced defense response by lowering expression of genes involved in defense response, suggesting a role of N and P nutrition in plant disease resistance. In conclusion, we provide detailed information on how LN, LP, and LNP affect different processes in soybean roots on the molecular and physiological levels.

Identification of key genes associated with secondary metabolites biosynthesis by system network analysis in Valeriana officinalis.

Valeriana officinalis is a medicinal plant, a source of bioactive chemical compounds and secondary metabolites which are applied in pharmaceutical industries. The advent of ethnomedicine has provided alternatives for disease treatment and has increased demands for natural products and bioactive compounds. A set of preliminary steps to answers for such demands can include integrative omics for systems metabolic engineering, as an approach that contributes to the understanding of cellular metabolic status. There is a growing trend of this approach for genetically engineering metabolic pathways in plant systems, by which natural and synthetic compounds can be produced. As in the case of most medicinal plants, there are no sufficient information about molecular mechanisms involved in the regulation of metabolic pathways in V. officinalis. In this research, systems biology was performed on the RNA-seq transcriptome and metabolome data to find key genes that contribute to the synthesis of major secondary metabolites in V. officinalis. The R Package Weighted Gene Co-Expression Network Analysis (WGCNA) was employed to analyze the data. Based on the results, some major modules and hub genes were identified to be associated with the valuable secondary metabolites. In addition, some TF-encoding genes, including AP2/ERF-ERF, WRKY and NAC TF families, as well as some regulatory factors including protein kinases and transporters were identified. The results showed that several novel hub genes, such as PCMP-H24, RPS24B, ANX1 and PXL1, may play crucial roles in metabolic pathways. The current findings provide an overall insight into the metabolic pathways of V. officinalis and can expand the potential for engineering genome-scale pathways and systems metabolic engineering to increase the production of bioactive compounds by plants.

Prepubertal exposure to high dose of cadmium induces hypothalamic injury through transcriptome profiling alteration and neuronal degeneration in female rats.

Cadmium (Cd) is a toxic metal, which seems to be crucial during the prepubertal period. Cd can destroy the structural integrity of the blood-brain barrier (BBB) and enters into the brain. Although the brain is susceptible to neurotoxicity induced by Cd, the effects of Cd on the brain, particularly hypothalamic transcriptome, are still relatively poorly understood. Therefore, we investigated the molecular effects of Cd exposure on the hypothalamus by profiling the transcriptomic response of the hypothalamus to high dose of Cd (25 mg/kg bw/day cadmium chloride (CdCl2)) during the prepubertal period in Sprague-Dawley female rats. After sequencing and annotation, differential expression analysis revealed 1656 genes that were differentially expressed that 108 of them were classified into 37 transcription factor (TF) families. According to gene ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, these differentially expressed genes (DEGs) were involved in different biological processes and neurological disorders including Alzheimer's disease (AD), Huntington's disease (HD), and Parkinson's disease (PD), prolactin signaling pathway, PI3K/Akt signaling, and dopaminergic synapse. Five transcripts were selected for further analyses with Real-time quantitative PCR (RT-qPCR). The RT-qPCR results were mostly consistent with those from the high throughput RNA sequencing (RNA-seq). Cresyl violet staining clearly showed an increased neuronal degeneration in the dorsomedial hypothalamus (DMH) and arcuate (Arc) nuclei of the CdCl2 group. Overall, this study demonstrates that prepubertal exposure to high doses of Cd induces hypothalamic injury through transcriptome profiling alteration in female rats, which reveals the new mechanisms of pathogenesis of Cd in the hypothalamus.

Tissue-specific transcriptional biomarkers in medicinal plants: Application of large-scale meta-analysis and computational systems biology.

Biosynthesis of secondary metabolites in plant is a complex process, regulated by many genes and influenced by several factors. In recent years, the next-generation sequencing (NGS) technology and advanced statistical analysis such as meta-analysis and computational systems biology have provided novel opportunities to overcome biological complexity. Here, we performed a meta-analysis on publicly available transcriptome datasets of twelve economically significant medicinal plants to identify differentially expressed genes (DEGs) between shoot and root tissues and to find the key molecular features which may be effective in the biosynthesis of secondary metabolites. Meta-analysis identified a total of 880 genes with differential expression between two tissues. Functional enrichment and KEGG pathway analysis indicated that the functions of those DEGs are highly associated with the developmental process, starch metabolic process, response to stimulus, porphyrin and chlorophyll metabolism, biosynthesis of secondary metabolites and phenylalanine metabolism. In addition, systems biology analysis of the DEGs was applied to find protein-protein interaction network and discovery of significant modules. The detected modules were associated with hormone signal transduction, transcription repressor activity, response to light stimulus and epigenetic processes. Finally, analysis was extended to search for putative miRNAs that are associated with DEGs. A total of 31 miRNAs were detected which belonged to 16 conserved families. The present study provides a comprehensive view to better understand the tissue-specific expression of genes and mechanisms involved in secondary metabolites synthesis and may provide candidate genes for future researches to improve yield of secondary metabolites.