De Novo RNA Sequencing and Transcriptome Analysis of Colletotrichum gloeosporioides ES026 Reveal Genes Related to Biosynthesis of Huperzine A

Abstract

Huperzine A is important in the treatment of Alzheimer’s disease. There are major challenges for the mass production of huperzine A from plants due to the limited number of huperzine-A-producing plants, as well as the low content of huperzine A in these plants. Various endophytic fungi produce huperzine A. Colletotrichum gloeosporioides ES026 was previously isolated from a huperzine-A-producing plant Huperzia serrata, and this fungus also produces huperzine A. In this study, de novo RNA sequencing of C. gloeosporioides ES026 was carried out with an Illumina HiSeq2000. A total of 4,324,299,051 bp from 50,442,617 high-quality sequence reads of ES026 were obtained. These raw data were assembled into 24,998 unigenes, 40,536,684 residues and 19,790 genes. The majority of the unique sequences were assigned to corresponding putative functions based on BLAST searches of public databases. The molecular functions, biological processes and biochemical pathways of these unique sequences were determined using gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) assignments. A gene encoding copper amine oxidase (CAO) (unigene 9322) was annotated for the conversion of cadaverine to 5-aminopentanal in the biosynthesis of huperzine A. This gene was also detected in the root, stem and leaf of H. serrata. Furthermore, a close relationship was observed between expression of the CAO gene (unigene 9322) and quantity of crude huperzine A extracted from ES026. Therefore, CAO might be involved in the biosynthesis of huperzine A and it most likely plays a key role in regulating the content of huperzine A in ES026.

Endophytic fungi residing in medicinal plants have the ability to produce the same or similar pharmacologically active secondary metabolites as their hosts

“…Similar fi ndings have been reported in the case of huperzine A producing endophytes. Their fungal amine oxidase genes have been found to present low similarities to the corresponding plant genes, and only conserved consensus sequences were present by the fungal and plant functional amine oxidase proteins Zhang et al, 2015), which supports the co-evolution theory rather than the HGT theory. This has been well established in the case of gibberellin biosynthetic pathways in fungi and higher plants where diff erences in genes and enzymes indicated converged evolution of GA metabolic pathways (Böm-ke and Tudzynski, 2009).…”

Transcriptome Analysis Reveals the Genetic Basis of the Resveratrol Biosynthesis Pathway in an Endophytic Fungus (Alternaria sp. MG1) Isolated from Vitis vinifera

“…As a newly explored biological resource, endophytic fungi offer an excellent opportunity to produce active compounds without the same limitations as plant resources. However, critical obstacles such as low and unstable production of metabolites greatly inhibit the application of these fungi (Zhang et al, 2015 ). Understanding the biosynthesis pathway and relative key genes is essential for an intensive control of their viability and stability.…”

“…Currently, transcriptome sequencing is an effective approach for mining genetic information from unknown genome sequences, and is useful in gene sequencing, new gene discovery, and studying gene regulation (Wang et al, 2009 ). This method can produce millions of short cDNA reads and provide useful information on gene expression, based on RNA abundance (Peng et al, 2014 ; Zhang et al, 2015 ). Next-generation sequencing technologies such as Roche 454 and Illumina RNA-Seq have emerged during a golden age of scientific discovery (Margulies et al, 2005 ; Metzker, 2010 ).…”

“…In recent years, a newly developed platform (Illumina Hiseq 2000/2500) has been successfully used in de novo sequencing experiments, providing a new transcriptome sequencing approach for non-model species, especially fungal species, without a reference genome (Zhang et al, 2013a ; Liu et al, 2014 ; Lou et al, 2014 ). This platform can produce a large amount of genetic information for many analysis types, such as comparative transcriptome analysis (Quan et al, 2015 ; Qiu et al, 2016 ), SSR identification (Yu et al, 2014 ; Huang et al, 2015 ) and uncovering unknown biosynthesis/metabolism pathways, in particular (Zhang et al, 2015 ; Zondag et al, 2016 ). Compared with the 454 technology and other sequencing platforms, the Illumina platform has optimized the traditional sequencing method, and is advantageous in its high throughput, high sensitivity, high accuracy and low cost, distinguishing it from its competitors.…”

Differences in huperzine A yield when different amino acids were added for in vitro thallus culture of Huperzia serrata (Thunb)Trev and gene expression analysis

“…Subsequently, they obtained RNA from this strain for sequencing analysis and individually validated the expression of copper amine oxidase (CAO) in the roots, stems, and leaves of H. serrata. They found that CAO may participate in HupA biosynthesis and may play a crucial role in regulating HupA yield in ES026 [9] . In 2017, Yang et al reported the transcriptome analysis of four different H. serrata tissues (roots, stems, leaves, and sporangias) and identified LDC, CAO, and PKS in transcripts [10] .…”