A novel Myb transcription factor represses conidiation and cephalosporin production in Acremonium chrysogenum
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ABSTRACT:
PROVIDER: PRJNA419940 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR6325100_1.fastq.gz | Fastqsanger.gz | |||
| SRR6325100_2.fastq.gz | Fastqsanger.gz | |||
| SRR6325101_1.fastq.gz | Fastqsanger.gz | |||
| SRR6325101_2.fastq.gz | Fastqsanger.gz | |||
| SRR6325102_1.fastq.gz | Fastqsanger.gz |
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Project description:Acremonium chrysogenum is the industrial producer of cephalosporin C. We isolated a mutant (AC554) from a T-DNA inserted library of A. chrysogenum. AC554 exhibited reduced conidiation and lack of cephalosporin C production. In consistent, the transcription of cephalosporin biosynthetic genes pcbC and cefEF was obviously decreased in AC554. TAIL-PCR and sequence analysis indicated that a T-DNA was inserted in the upstream of an open reading frame (ORF) which was designated AcmybA. Sequence analysis indicated that AcmybA encodes a novel Myb domain containing transcriptional factor. Observation of red fluorescence protein (RFP) tagged AcMybA showed that AcMybA is naturally located in the nuclear of A. chrysogenum. Transcription analysis demonstrated that AcmybA was overexpressed in AC554. In contrast with AC554, the AcmybA deleted mutant (DAcmybA) overproduced conidia in LPE medium and increased cephalosporin production during fermentation. To determine the genes under the influence of AcmybA, we sequenced and compared the transcriptome of DAcmybA, AC554 and the wild-type strain at different developmental stages. Results confirmed the repression of AcMybA on the key conidiation regulatory gene AcbrlA and the cephalosporin biosynthetic genes. Among the targets of AcMybA, 10 putative regulatory genes were selected and overexpressed in A. chrysogenum. Taken together, our results indicate that AcMybA negatively regulates conidiation and cephalosporin production in A. chrysogenum.
2017-11-28 | GSE107388 | GEO
Project description:Penicillium chrysogenum was successfully engineered to produce a novel carbamoylated cephalosporin that can be used as a synthon for semi-synthetic cephalosporins. To this end, structural genes for Acremonium chrysogenum expandase/hydroxylase and Streptomyces clavuligerus carbamoyltransferase were expressed in a penicillinG high-producing strain of P. chrysogenum. Growth of the engineered strain in the presence of the side-chain precursor adipic acid resulted in production of adipoyl-7-amino-3-carbamoyloxymethyl-3-cephem-4-carboxylic acid (ad7-ACCCA) and of several adipoylated pathway intermediates. A combinatorial chemostat-based transcriptome study, in which the ad7-ACCCA- producing strain and a strain lacking key genes in β-lactam synthesis were grown in the presence and absence of adipic acid, enabled the dissection of transcriptional responses to adipic acid per se and to ad7-ACCCA production. In chemostat cultures of both strains, adipic acid served as an additional carbon source. Transcriptome analysis supported an earlier proposal, based on 13C-labelling studies, that adipic acid catabolism in P. chrysogenum occurs via β-oxidation and enabled the identification of putative genes for enzymes involved in mitochondrial and peroxisomal β-oxidation pathways. Several of the genes that showed a specifically altered transcript level in ad7-ACCCA-producing cultures were previously implicated in oxidative stress responses. As strain improvement programmes lead to increased specific productivity and yields, a deeper understanding of these stress responses is likely to be important to also achieve high ad7-ACCCA titers with engineered strains of P. chrysogenum.
2009-01-05 | GSE12612 | GEO
Project description:Genome sequence of Acremonium chrysogenum, the producer of the beta-lactam antibiotic cephalosporin C
| PRJNA248608 | ENA
Project description:Production of cephalosporin precursors with recombinant strains of Penicillium chrysogenum has improved the economics and reduced the environmental impact of industrial cephalosporin production. The engineered P. chrysogenum strains used in these processes express heterologous enzymes that convert the intermediate acyl-6-aminopenicillanic acid into different tailor-made compounds. Activation of the cephalosporin side-chain precursor to its corresponding CoA thioester is an essential step for its incorporation into the β-lactam backbone. To identify the acyl-CoA ligase involved in activation of adipic acid, a frequently used cephalosporin side-chain precursor, we searched the genome of P.chrysogenum for putative structural genes encoding acyl-CoA ligases. Chemostat-based transcriptome analysis was then used to identify the one presenting the highest expression level when cells were grown in the presence of adipic acid. Deletion of the gene renamed aclA, led to a 32% decreased specific rate of adipic acid consumption and a three-fold reduction of adipoyl-6-aminopenicillanic acid levels in chemostat cultures of P. chrysogenum, but did not affect penicillin production. After cloning the gene and overexpressing it in Escherichia coli, its purified protein product was shown to have adipoyl-CoA ligase, but no phenylacetyl-CoA ligtase activity. Finally, by fusing the gene to a sequence encoding cyan fluorescent protein, the resulting fusion protein localized to microbodies, which indicates that activation of the side-chain precursor adipic acid takes place in this compartment, where also the subsequent acyltransferase step takes place. Identification and functional characterization of this adipoyl-CoA ligtase gene may aid in developing future metabolic engineering strategies for improving the production of different cephalosporins.
2009-08-01 | GSE12617 | GEO
Project description:In filamentous fungi, secondary metabolism is often linked with developmental processes such as conidiation. In this study we analyzed the link between secondary metabolism and conidiation in the main industrial producer of the β-lactam antibiotic penicillin, the ascomycete Penicillium chrysogenum. Therefore, we generated mutants defective in two central regulators of conidiation, the transcription factors BrlA and StuA, respectively. Inactivation of both BrlA and StuA blocked conidiation and altered hyphal morphology during growth on solid media, as shown by light and scanning electron microscopy, but did not affect biomass production during liquid submerged growth. Genome-wide transcriptional profiling identified a complex StuA- and BrlA-dependent regulatory network, including genes previously shown to be involved in development and secondary metabolism. Remarkably, inactivation of StuA, but not BrlA, drastically down-regulated expression of the penicillin biosynthetic gene cluster during solid and liquid submerged growth. In agreement, penicillin V production was wild type-like in BrlA-deficient strains but 99 % decreased in StuA-deficient strains during liquid submerged growth as shown by HPLC analysis. Thus, among identified regulators of penicillin V production StuA has the most severe influence. Over-expression of StuA increased the transcript levels of BrlA and AbaA (another developmental regulator), de-repressed conidiation during liquid submerged growth, but did not affect penicillin V productivity. Taken together, these data demonstrate an intimate but not exclusive link between regulation of development and secondary metabolism in P. chrysogenum. Transcriptomes of PcbrlA- and PcstuA- deletion mutants were compared with expression data from recipient strain deltaPcku70 as a control Mycelia from the transformants and the reference strain were harvested at successive stages of development for RNA extraction and hybridization on Affymetrix microarrays.
2011-01-04 | E-GEOD-23326 | biostudies-arrayexpress
Project description:In filamentous fungi, secondary metabolism is often linked with developmental processes such as conidiation. In this study we analyzed the link between secondary metabolism and conidiation in the main industrial producer of the β-lactam antibiotic penicillin, the ascomycete Penicillium chrysogenum. Therefore, we generated mutants defective in two central regulators of conidiation, the transcription factors BrlA and StuA, respectively. Inactivation of both BrlA and StuA blocked conidiation and altered hyphal morphology during growth on solid media, as shown by light and scanning electron microscopy, but did not affect biomass production during liquid submerged growth. Genome-wide transcriptional profiling identified a complex StuA- and BrlA-dependent regulatory network, including genes previously shown to be involved in development and secondary metabolism. Remarkably, inactivation of StuA, but not BrlA, drastically down-regulated expression of the penicillin biosynthetic gene cluster during solid and liquid submerged growth. In agreement, penicillin V production was wild type-like in BrlA-deficient strains but 99 % decreased in StuA-deficient strains during liquid submerged growth as shown by HPLC analysis. Thus, among identified regulators of penicillin V production StuA has the most severe influence. Over-expression of StuA increased the transcript levels of BrlA and AbaA (another developmental regulator), de-repressed conidiation during liquid submerged growth, but did not affect penicillin V productivity. Taken together, these data demonstrate an intimate but not exclusive link between regulation of development and secondary metabolism in P. chrysogenum. Transcriptomes of PcbrlA- and PcstuA- deletion mutants were compared with expression data from recipient strain deltaPcku70 as a control
2011-01-04 | GSE23326 | GEO
Project description:Earlier we have shown important roles of MYB in pancreatic tumor pathobiology. To better understand the role of MYB in the tumor microenvironment and identify MYB-associated secreted biomarker proteins, we conducted mass spectrometry analysis of the secretome from MYB-modulated and control pancreatic cancer cell lines. We also performed in silico analyses to determine MYB-associated biofunctions, gene networks and altered biological pathways. We further investigated the secreted proteins for their potential biomarker properties.
2021-09-08 | PXD015126 | Pride
Project description:Background: Microbial gene expression is to a large extend determined by environmental growth conditions. Differential gene expression analysis between two conditions has been frequently used to reveal regulatory networks and to assign physiological function to unknown genes. In nature, microorganisms cohabit however these interactions have been rarely studied and reproduced in laboratory set-up. Thus to quantitatively explore the genome-wide responses of microbial interaction, we co-cultivated Penicillium chrysogenum and Bacillus subtilis in chemostat culture. Results: Time course expression analysis of P. chrysogenum to co-cultivation with B. subtilis was carried out to understand the natural responses of P. chrysogenum to prokaryotes. Steady state chemostats of P. chrysogenum in non-B-lactam producing conditions was pulsed with B. subtilis and co-cultivation was followed for 72 hours. The dynamic physiological and transcriptional responses of P. chrysogenum in mixed culture were monitored. B. subtilis outcompeted growth of P. chrysogenum resulting in an increased B. subtilis biomass by more than three fold of its original size and a reduction in P. chrysogenum biomass to half of its original size after 72 h of mixed culture. Genes of the penicillin pathway, synthesis of the side-chain and precursors were overall unresponsive to the presence of B. subtilis. Moreover Penicillium polyketide synthase and nonribosomal peptide synthetase genes either remained silent or down-regulated, whereas genes responsible for protein synthesis, metabolism, energy conservation, respiration and transport were upregulated in the presence of B. subtilis. Among highly responsive genes, two putative B-1,3 endoglucanase (mutanase) genes viz Pc12g07500 and Pc12g13330 were upregulated by more than 15-fold and 8-fold respectively. Measurement of enzyme activity in the supernatant of mixed culture confirmed that the co-cultivation with B. subtilis induced mutanase production in P. chrysogenum. Mutanase activity was not observed in pure cultures of P. chrysogenum and B. subtilis or when P. chrysogenum was co-cultured with B. subtilis supernatant or heat inactivated B. subtilis cells. However, mutanase production was observed in cultures of P. chrysogenum pulsed with filter sterilized supernatants from mixed cultures P. chrysogenum and B. subtilis. Heterologous expression of Pc12g07500 and Pc12g13330 genes in Saccharomyces cerevisiae confirmed that at least Pc12g07500 encoded an B-1,3 endoglucanase. Conclusion: Time course transcriptional profiling of P. chrysogenum revealed several differentially expressed genes during mixed culture, potentially reflecting interactions between the eukaryotic and the prokaryotic systems. -1,3 endoglucanase produced by P. chrysogenum against B. subtilis signals may have application in improving the efficacy of antibiotics by degrading exopolysacchride biofilms of pathogenic bacteria.
2014-05-12 | GSE53286 | GEO