Chaetoglobosin A(cheA)is a complex indole alkaloid exhibiting preferential cytotoxicity against plant patho-gens,parasites,and tumor cells.However,the limited production and high synthesis costs of cheA impede its wid...Chaetoglobosin A(cheA)is a complex indole alkaloid exhibiting preferential cytotoxicity against plant patho-gens,parasites,and tumor cells.However,the limited production and high synthesis costs of cheA impede its widespread application.Tryptophan serves as a precursor for cheA biosynthesis,and strategic modification of the expression of key genes represents a novel approach to enhance the target yield.Herein,CgAS,a gene encoding anthranilic acid synthase involved in tryptophan synthesis,was identified through bioinformatics analysis and overexpressed via a promoter optimization strategy in Chaetomium globosum W7.The AS1 and AS3 mutants,in which the CgAS gene was constitutively overexpressed under the control of promoter oliC,presented a significant increase in tryptophan accumulation.CgAS overexpression caused a dramatic increase in cheA production,reaching a maximum yield of 217.81 mg/L during the stationary phase,which was 3.73-fold higher than that noted in the wild-type strain.Interestingly,AS1 and AS3 mutants exhibited a substantial upregulation in the transcription levels of critical genes involved in cheA biosynthesis.Phenotypic characterization and metabolomic analysis indicated that tryptophan accumulation strengthened microbial nitrogen metabolism,which not only provided sufficient precursors for secondary metabolism,but also functioned as an essential energy source to accelerate fungal development and sporulation.These findings illustrate the impact of precursor accumulation on indole alkaloid biosynthesis and provide novel insights for optimizing the production of biopesticides and clinical drugs.展开更多
Chaetomium globosum is one of the most common fungi in nature. It is best known for producing chaetoglobosins; however, the molecular basis of chaetoglobosin biosynthesis is poorly understood in this fungus. In this s...Chaetomium globosum is one of the most common fungi in nature. It is best known for producing chaetoglobosins; however, the molecular basis of chaetoglobosin biosynthesis is poorly understood in this fungus. In this study, we utilized RNA inter- ference （RNAi） to characterize a polyketide synthase gene, pks-1, in C. globosum that is involved in the production of chaeto- globosin A. When pks-1 was knocked down by RNAi, the production of chaetoglobosin A dramatically decreased. Knock-down mutants also displayed a pigment-deficient phenotype. These results suggest that the two polyketides, melanin and chaetoglobosin, are likely to share common biosynthetic steps. Most importantly, we found that pks-I also plays a critical role in sporulation. The silenced mutants ofpks-1 lost the ability to produce spores. We propose that polyketides may modulate cellular development via an unidentified action. We also suggest that C. globosum pks-1 is unique because of its triple role in melanin formation, chaetoglobosin biosynthesis and sporulation. This work may shed light on chaetoglobosin biosynthesis and indicates a relationship between secondary metabolism and fungal morphogenesis.展开更多
Cytochalasans,with diverse structures and pharmacological activities,are a class of compounds containing isoindolinone moieties fused to the tricyclic or tetracyclic ring system.Chaetoglobosin A(cheA),mainly produced ...Cytochalasans,with diverse structures and pharmacological activities,are a class of compounds containing isoindolinone moieties fused to the tricyclic or tetracyclic ring system.Chaetoglobosin A(cheA),mainly produced by Chaetomium globosum,is the most abundant cytochalasan.However,limited understanding of transcriptional regulation of morphological development and cheA biosynthesis in C.globosum has hindered cheA application in agriculture and biomedical field.This study examined the regulatory role of CgVeA gene in C.globosum.CgVeA had significant effect on secondary metabolites production in C.globosum,similar to that reported in other filamentous fungi.Inactivation of CgVeA caused an obvious decrease in cheA production from 51.32 to 19.76 mg/L under dark conditions.In contrast,CgVeA overexpression resulted in a dramatic increase in cheA production,reaching 206.59 mg/L under light conditions,which was higher than that noted under dark condition.The RT-qPCR results confirmed that CgVeA,as a light responsive regulator,positively regulated cheA biosynthesis by controlling the expression of core genes of the cheA biosynthetic gene cluster and other relevant regulators.Electrophoretic mobility shift assays proved that CgVeA directly regulated LaeA,cheR,and p450,and indirectly regulated PKS.Moreover,CgVeA had a significant effect on the regulation of asexual spores production.When compared with wild-type C.globosum,CgVeA-silenced and CgVeA overexpression mutants presented remarkable differences in sporulation,irrespective of light or dark condition.Besides,CgVeA expression was speculated to negatively regulate spore formation.These findings illustrated the regulatory mechanism of a hypothetical global regulator,CgVeA,in C.globosum,suggesting its potential application in industrial-scale cheA biosynthesis.展开更多
基金supported by the Key Research and Development Program of Hainan Province(ZDYF2024SHFZ046)the Fundamental Research Funds for the Central Universities(226-2024-00019)+2 种基金the Na-tional Natural Science Foundation of Zhejiang Province(LDT23D06022D06)the Project of the Donghai Laboratory(Z24ZJ004P)the Science Foundation of Donghai Laboratory(L24QH014).
文摘Chaetoglobosin A(cheA)is a complex indole alkaloid exhibiting preferential cytotoxicity against plant patho-gens,parasites,and tumor cells.However,the limited production and high synthesis costs of cheA impede its widespread application.Tryptophan serves as a precursor for cheA biosynthesis,and strategic modification of the expression of key genes represents a novel approach to enhance the target yield.Herein,CgAS,a gene encoding anthranilic acid synthase involved in tryptophan synthesis,was identified through bioinformatics analysis and overexpressed via a promoter optimization strategy in Chaetomium globosum W7.The AS1 and AS3 mutants,in which the CgAS gene was constitutively overexpressed under the control of promoter oliC,presented a significant increase in tryptophan accumulation.CgAS overexpression caused a dramatic increase in cheA production,reaching a maximum yield of 217.81 mg/L during the stationary phase,which was 3.73-fold higher than that noted in the wild-type strain.Interestingly,AS1 and AS3 mutants exhibited a substantial upregulation in the transcription levels of critical genes involved in cheA biosynthesis.Phenotypic characterization and metabolomic analysis indicated that tryptophan accumulation strengthened microbial nitrogen metabolism,which not only provided sufficient precursors for secondary metabolism,but also functioned as an essential energy source to accelerate fungal development and sporulation.These findings illustrate the impact of precursor accumulation on indole alkaloid biosynthesis and provide novel insights for optimizing the production of biopesticides and clinical drugs.
基金the National Natural Science Foundation of China (Grant No. 30970084)the National Basic Research Program of China (Grant No. 2007CB707801)
文摘Chaetomium globosum is one of the most common fungi in nature. It is best known for producing chaetoglobosins; however, the molecular basis of chaetoglobosin biosynthesis is poorly understood in this fungus. In this study, we utilized RNA inter- ference （RNAi） to characterize a polyketide synthase gene, pks-1, in C. globosum that is involved in the production of chaeto- globosin A. When pks-1 was knocked down by RNAi, the production of chaetoglobosin A dramatically decreased. Knock-down mutants also displayed a pigment-deficient phenotype. These results suggest that the two polyketides, melanin and chaetoglobosin, are likely to share common biosynthetic steps. Most importantly, we found that pks-I also plays a critical role in sporulation. The silenced mutants ofpks-1 lost the ability to produce spores. We propose that polyketides may modulate cellular development via an unidentified action. We also suggest that C. globosum pks-1 is unique because of its triple role in melanin formation, chaetoglobosin biosynthesis and sporulation. This work may shed light on chaetoglobosin biosynthesis and indicates a relationship between secondary metabolism and fungal morphogenesis.
基金This work was supported in part by grant from the Harbin Science and Technology Project(No.2016AB3AP042).
文摘Cytochalasans,with diverse structures and pharmacological activities,are a class of compounds containing isoindolinone moieties fused to the tricyclic or tetracyclic ring system.Chaetoglobosin A(cheA),mainly produced by Chaetomium globosum,is the most abundant cytochalasan.However,limited understanding of transcriptional regulation of morphological development and cheA biosynthesis in C.globosum has hindered cheA application in agriculture and biomedical field.This study examined the regulatory role of CgVeA gene in C.globosum.CgVeA had significant effect on secondary metabolites production in C.globosum,similar to that reported in other filamentous fungi.Inactivation of CgVeA caused an obvious decrease in cheA production from 51.32 to 19.76 mg/L under dark conditions.In contrast,CgVeA overexpression resulted in a dramatic increase in cheA production,reaching 206.59 mg/L under light conditions,which was higher than that noted under dark condition.The RT-qPCR results confirmed that CgVeA,as a light responsive regulator,positively regulated cheA biosynthesis by controlling the expression of core genes of the cheA biosynthetic gene cluster and other relevant regulators.Electrophoretic mobility shift assays proved that CgVeA directly regulated LaeA,cheR,and p450,and indirectly regulated PKS.Moreover,CgVeA had a significant effect on the regulation of asexual spores production.When compared with wild-type C.globosum,CgVeA-silenced and CgVeA overexpression mutants presented remarkable differences in sporulation,irrespective of light or dark condition.Besides,CgVeA expression was speculated to negatively regulate spore formation.These findings illustrated the regulatory mechanism of a hypothetical global regulator,CgVeA,in C.globosum,suggesting its potential application in industrial-scale cheA biosynthesis.
