The parasitic fungus Claviceps purpurea has been used for decades by the pharmaceutical industry as a valuable producer of ergot alkaloids. As the biosynthetic pathway of ergot alkaloids involves a common precursor L-tryptophan, targeted genetic modification of the related genes may improve production yield. In this work, the S76L mutated version of the trpE gene encoding anthranilate synthase was constitutively overexpressed in the fungus with the aim of overcoming feedback inhibition of the native enzyme by an excess of tryptophan. In another approach, the dmaW gene encoding dimethylallyltryptophan synthase, which produces a key intermediate for the biosynthesis of ergot alkaloids, was also constitutively overexpressed. Each of the above manipulations led to a significant increase (up to 7-fold) in the production of ergot alkaloids in submerged cultures.
The fungus Claviceps purpurea is a biotrophic phytopathogen widely used in the pharmaceutical industry for its ability to produce ergot alkaloids (EAs). The fungus attacks unfertilized ovaries of grasses and forms sclerotia, which represent the only type of tissue where the synthesis of EAs occurs. The biosynthetic pathway of EAs has been extensively studied; however, little is known concerning its regulation. Here, we present the quantitative transcriptome analysis of the sclerotial and mycelial tissues providing a comprehensive view of transcriptional differences between the tissues that produce EAs and those that do not produce EAs and the pathogenic and non-pathogenic lifestyle. The results indicate metabolic changes coupled with sclerotial differentiation, which are likely needed as initiation factors for EA biosynthesis. One of the promising factors seems to be oxidative stress. Here, we focus on the identification of putative transcription factors and regulators involved in sclerotial differentiation, which might be involved in EA biosynthesis. To shed more light on the regulation of EA composition, whole transcriptome analysis of four industrial strains differing in their alkaloid spectra was performed. The results support the hypothesis proposing the composition of the amino acid pool in sclerotia to be an important factor regulating the final structure of the ergopeptines produced by Claviceps purpurea.
- MeSH
- biotechnologie MeSH
- Claviceps genetika metabolismus MeSH
- exprese genu MeSH
- fungální proteiny genetika metabolismus MeSH
- geny hub MeSH
- jednonukleotidový polymorfismus MeSH
- námelové alkaloidy biosyntéza MeSH
- oxidační stres MeSH
- peptidsynthasy genetika metabolismus MeSH
- průmyslová mikrobiologie MeSH
- sekvence aminokyselin MeSH
- sekvenční homologie aminokyselin MeSH
- stanovení celkové genové exprese MeSH
- Publikační typ
- časopisecké články MeSH
Disease symptoms of some phytopathogenic fungi are associated with changes in cytokinin (CK) levels. Here, we show that the CK profile of ergot-infected rye plants is also altered, although no pronounced changes occur in the expression of the host plant's CK biosynthesis genes. Instead, we demonstrate a clearly different mechanism: we report on the first fungal de novo CK biosynthesis genes, prove their functions and constitute a biosynthetic pathway. The ergot fungus Claviceps purpurea produces substantial quantities of CKs in culture and, like plants, expresses enzymes containing the isopentenyltransferase and lonely guy domains necessary for de novo isopentenyladenine production. Uniquely, two of these domains are combined in one bifunctional enzyme, CpIPT-LOG, depicting a novel and potent mechanism for CK production. The fungus also forms trans-zeatin, a reaction catalysed by a CK-specific cytochrome P450 monooxygenase, which is encoded by cpp450 forming a small cluster with cpipt-log. Deletion of cpipt-log and cpp450 did not affect virulence of the fungus, but Δcpp450 mutants exhibit a hyper-sporulating phenotype, implying that CKs are environmental factors influencing fungal development.
- MeSH
- alkyltransferasy a aryltransferasy metabolismus MeSH
- Claviceps genetika růst a vývoj metabolismus MeSH
- cytokininy biosyntéza MeSH
- delece genu MeSH
- geny hub genetika MeSH
- isopentenyladenosin biosyntéza MeSH
- rostlinné geny genetika MeSH
- systém (enzymů) cytochromů P-450 genetika MeSH
- žito mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Ergot alkaloids produced by the fungus Claviceps parasitizing on cereals, include three major groups: clavine alkaloids, d-lysergic acid and its derivatives and ergopeptines. These alkaloids are important substances for the pharmatech industry, where they are used for production of anti-migraine drugs, uterotonics, prolactin inhibitors, anti-Parkinson agents, etc. Production of ergot alkaloids is based either on traditional field cultivation of ergot-infected rye or on submerged cultures of the fungus in industrial fermentation plants. In 2010, the total production of these alkaloids in the world was about 20,000 kg, of which field cultivation contributed about 50%. This review covers the recent advances in understanding of the genetics and regulation of biosynthesis of ergot alkaloids, focusing on possible applications of the new knowledge to improve the production yield.
- MeSH
- Claviceps genetika metabolismus MeSH
- enzymy metabolismus MeSH
- fermentace MeSH
- genetické inženýrství metody MeSH
- molekulární struktura MeSH
- námelové alkaloidy biosyntéza chemie genetika farmakologie MeSH
- průmyslová mikrobiologie metody MeSH
- žito mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
