In filamentous fungi,nitrogen metabolism is repressed by GATA-type zinc finger transcription factors.Nitrogen metabolite repression has been found to affect antibiotic production,but the mechanism is still poorly unde...In filamentous fungi,nitrogen metabolism is repressed by GATA-type zinc finger transcription factors.Nitrogen metabolite repression has been found to affect antibiotic production,but the mechanism is still poorly understood.AcareB,encoding a homologue of fungal GATA-type regulatory protein,was cloned from Acremonium chrysogenum.Gene disruption and genetic complementation demonstrated that AcareB plays a key role in utilization of ammonium,glutamine and urea.In addition,significant reduction of cephalosporin production in the AcareB disruption mutant indicated that AcareB is important for cephalosporin production.In consistence with it,the transcriptional level of cephalosporin biosynthetic genes was significantly decreased in the AcareB disruption mutant.Electrophoretic mobility shift assay showed that AcAREB directly bound to the intergenic regions of pcbAB-pcbC,cefD1-cefD2 and cefEF-cefG.Sequence analysis showed that all the AcAREB binding sites contained the consensus GATA elements.AcareB is negatively autoregulated during cephalosporin production.Moreover,another GATA zinc-finger protein encoded by AcareA positively regulates the transcription of AcareB.However,AcareB does not regulate the transcription of AcareA.These results indicated that AcAREB plays an important role in both regulation of nitrogen metabolism and cephalosporin production in A.chrysogenum.展开更多
Acremonium chrysogenum is an important filamentous fungus which produces cephalosporin C in industry.This review summarized the study on genetic engineering of Acremonium chrysogenum,including biosynthesis and regulat...Acremonium chrysogenum is an important filamentous fungus which produces cephalosporin C in industry.This review summarized the study on genetic engineering of Acremonium chrysogenum,including biosynthesis and regulation for fermentation of cephalosporin C,molecular techniques,molecular breeding and transcriptomics of Acremonium chrysogenum.We believe with all the techniques available and full genomic sequence,the industrial strain of Acremonium chrysogenum can be genetically modified to better serve the pharmaceutical industry.展开更多
Two new compounds,(22E)-25-carboxy-8β,14β-epoxy-4α,5α-dihydroxyergosta-2,22-dien-7-one(1) and fusidione(3), along with two known compounds, 5α,8α-epidioxy ergosta-6,22-diene-3β-ol(2) and microperfuranone(4), we...Two new compounds,(22E)-25-carboxy-8β,14β-epoxy-4α,5α-dihydroxyergosta-2,22-dien-7-one(1) and fusidione(3), along with two known compounds, 5α,8α-epidioxy ergosta-6,22-diene-3β-ol(2) and microperfuranone(4), were isolated from the fermentation products of the marine-sourced fungus Acremonium fusidioides RZ01. The structures of compounds 1 and 3 were elucidated by extensive spectroscopic methods, especially 2D NMR, and their absolute configurations were suggested on the basis of the circular dichroism spectral analysis and the NOESY data. Both new compounds showed inhibitory activity against HL-60 cells with IC50 values being16.6 and 44.9 μmol·L-1, respectively.展开更多
Acremonium species are prolific producers of therapeutic molecules which include the widely used beta-lactam antibiotic,cephalosporin.In light of their significant medical value,an efficient gene disruption method is ...Acremonium species are prolific producers of therapeutic molecules which include the widely used beta-lactam antibiotic,cephalosporin.In light of their significant medical value,an efficient gene disruption method is required for the physiological and biochemical studies on this genus of fungi.However,the number of selection markers that can be used for gene targeting is limited,which constrain the genetic analysis of multiple functional genes.In this study,we established a uridine auxotrophy based marker recycling system which achieves scarless gene deletion,and allows the use of the same selection marker in successive transformations in a deep seaderived fungus Acremonium sp.HDN16-126.We identified one homologue of Acremonium chrysogenum pyrG(also as a homologous gene of the yeast URA3)from HDN16-126,designated as pyrG-A1,which can be used as a selection marker on uridine free medium.We then removed pyrG-A1 from HDN16-126 genome via homologous recombination(HR)on MM medium with 5-fluoroortic acid(5-FOA),a chemical that can be converted into a toxin of 5-flurouracil by pyrG-A1 activity,thus generating the HDN16-126-△pyrG mutant strain which showed auxotrophy for uridine but insensitivity to 5-FOA and enabled the use of exogenous pyrG gene as both positive and negative selection marker to achieve the scarless deletion of target DNA fragments.We further applied this marker recycling system to successfully disrupt two target genes pepL(encodes a putative 2OG-Fe(Ⅱ)dioxygenase)and pepM(encodes a putative aldolase)identified from HDN16-126 genome,which are proposed to be functional genes related to 2-aminoisobutyric acid metabolism in fungi.This work is the first application of uridine auxotrophy based scarless gene deletion method in Acremonium species and shows promising potential in assisting sequential genetic analysis of filamentous fungi.展开更多
基金supported by the National Natural Science Foundation of China(31670091 and 31470177)
文摘In filamentous fungi,nitrogen metabolism is repressed by GATA-type zinc finger transcription factors.Nitrogen metabolite repression has been found to affect antibiotic production,but the mechanism is still poorly understood.AcareB,encoding a homologue of fungal GATA-type regulatory protein,was cloned from Acremonium chrysogenum.Gene disruption and genetic complementation demonstrated that AcareB plays a key role in utilization of ammonium,glutamine and urea.In addition,significant reduction of cephalosporin production in the AcareB disruption mutant indicated that AcareB is important for cephalosporin production.In consistence with it,the transcriptional level of cephalosporin biosynthetic genes was significantly decreased in the AcareB disruption mutant.Electrophoretic mobility shift assay showed that AcAREB directly bound to the intergenic regions of pcbAB-pcbC,cefD1-cefD2 and cefEF-cefG.Sequence analysis showed that all the AcAREB binding sites contained the consensus GATA elements.AcareB is negatively autoregulated during cephalosporin production.Moreover,another GATA zinc-finger protein encoded by AcareA positively regulates the transcription of AcareB.However,AcareB does not regulate the transcription of AcareA.These results indicated that AcAREB plays an important role in both regulation of nitrogen metabolism and cephalosporin production in A.chrysogenum.
文摘Acremonium chrysogenum is an important filamentous fungus which produces cephalosporin C in industry.This review summarized the study on genetic engineering of Acremonium chrysogenum,including biosynthesis and regulation for fermentation of cephalosporin C,molecular techniques,molecular breeding and transcriptomics of Acremonium chrysogenum.We believe with all the techniques available and full genomic sequence,the industrial strain of Acremonium chrysogenum can be genetically modified to better serve the pharmaceutical industry.
基金supported by the National Natural Science Foundation of China(No.81502951)
文摘Two new compounds,(22E)-25-carboxy-8β,14β-epoxy-4α,5α-dihydroxyergosta-2,22-dien-7-one(1) and fusidione(3), along with two known compounds, 5α,8α-epidioxy ergosta-6,22-diene-3β-ol(2) and microperfuranone(4), were isolated from the fermentation products of the marine-sourced fungus Acremonium fusidioides RZ01. The structures of compounds 1 and 3 were elucidated by extensive spectroscopic methods, especially 2D NMR, and their absolute configurations were suggested on the basis of the circular dichroism spectral analysis and the NOESY data. Both new compounds showed inhibitory activity against HL-60 cells with IC50 values being16.6 and 44.9 μmol·L-1, respectively.
基金funded by the National Key R&D Program of China(2019YFC0312504)the National Natural Science Foundation of China(41976105,81991522)+5 种基金Major Basic Research Programs of Natural Science Foundation of Shandong Province(ZR2019ZD18)the Fundamental Research Funds for the Central Universities(201941001)the NSFC-Shandong Joint Fund(U1906212)the Marine S&T Fund of Shandong Province for Pilot National Laboratory for Marine Science and Technology(Qingdao)(No.2018SDKJ0401-2)the Youth Innovation Plan of Shandong province(2019KJM004)the Taishan Scholar Youth Expert Program in Shandong Province(tsqn201812021).
文摘Acremonium species are prolific producers of therapeutic molecules which include the widely used beta-lactam antibiotic,cephalosporin.In light of their significant medical value,an efficient gene disruption method is required for the physiological and biochemical studies on this genus of fungi.However,the number of selection markers that can be used for gene targeting is limited,which constrain the genetic analysis of multiple functional genes.In this study,we established a uridine auxotrophy based marker recycling system which achieves scarless gene deletion,and allows the use of the same selection marker in successive transformations in a deep seaderived fungus Acremonium sp.HDN16-126.We identified one homologue of Acremonium chrysogenum pyrG(also as a homologous gene of the yeast URA3)from HDN16-126,designated as pyrG-A1,which can be used as a selection marker on uridine free medium.We then removed pyrG-A1 from HDN16-126 genome via homologous recombination(HR)on MM medium with 5-fluoroortic acid(5-FOA),a chemical that can be converted into a toxin of 5-flurouracil by pyrG-A1 activity,thus generating the HDN16-126-△pyrG mutant strain which showed auxotrophy for uridine but insensitivity to 5-FOA and enabled the use of exogenous pyrG gene as both positive and negative selection marker to achieve the scarless deletion of target DNA fragments.We further applied this marker recycling system to successfully disrupt two target genes pepL(encodes a putative 2OG-Fe(Ⅱ)dioxygenase)and pepM(encodes a putative aldolase)identified from HDN16-126 genome,which are proposed to be functional genes related to 2-aminoisobutyric acid metabolism in fungi.This work is the first application of uridine auxotrophy based scarless gene deletion method in Acremonium species and shows promising potential in assisting sequential genetic analysis of filamentous fungi.
