Newly identified Aspergillus nidulans PKA targets play a role in carbon-catabolite repression
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ABSTRACT:
ORGANISM(S): Aspergillus nidulans
PROVIDER: GSE116579 | GEO | 2018/07/04
REPOSITORIES: GEO
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Project description:Saprobic microorganisms, such as filamentous fungi of the genus Aspergillus, are of particular interest for biotechnological applications due to their natural capacity to secrete plant cell wall-degrading enzymes that are known as carbohydrate-active enzymes (CAZy). The presence of easily metabolizable sugars such as glucose, whose concentrations increase during plant biomass hydrolysis, results in the repression of CAZy-encoding genes, in a process known as carbon catabolite repression (CCR), which is undesired for the purpose of large-scale enzyme production. To date, the C2H2 transcription factor CreA has been described as the major CC-repressor in Aspergillus spp. Although CreA-mediated CCR has been extensively studied, less is known about the regulation of CreA itself and the role of post-translational modifications in this process. In this work, phosphorylation sites were identified by mass spectrometry on A. nidulans CreA and subsequently four sites, S262, S268, T308 and S319, were chosen to be mutated to non-phosphorylatable residues before their effects on CCR in both CC-repressing and de-repressing conditions were studied. Sites S262, S268 and T308 are important for CreA protein accumulation and cellular localization and repression of enzyme activities. In contrast, site S319 is key for CreA degradation and induction of enzyme activities. All sites were shown to be important for glycogen and trehalose metabolism. This study highlights the importance of CreA phosphorylation sites for the regulation of CCR. Furthermore, these sites are interesting targets for biotechnological strain engineering without the need to delete essential genes which can result in undesired side effects.
2021-02-18 | PXD018967 | Pride
Project description:A transcriptomic assessment of a kinase deletion mutant (ΔmpkA) and a control strain revealed many cellular activities this kinase is impacting. Studies were conducted in the model filamentous fungi, Aspergillus nidulans¸ under rich growth conditions. MpkA is a known kinase involved in the cell wall integrity signaling pathway. This pathway is activated upon cell wall stress and regulates production of cell wall related genes. Here, fungi were grown under non-stressful conditions. The comparison between the control strain and the deletion mutant transcript levels revealed previously unknown functions, such as iron starvation, that MpkA may be involved with.
2019-05-01 | GSE108325 | GEO
Project description:The ascomycete Trichoderma reesei is one of the most well studied cellulolytic fungi and widely used in the biotechnology industry, as in the production of second-generation bioethanol. Carbon catabolite repression (CCR) mechanism adopted by T. reesei is mediated by the transcription factor CRE1 and consists in the repression of genes related to the production of cellulase when a readily available carbon source is present in the medium. Using RNA sequencing this study aims to contribute to understanding of CCR during growth in cellulose and glucose, by comparing the mutant strain of T. reesei Δcre1 with its parental, QM9414.
2017-05-01 | GSE57374 | GEO
Project description:Plant biomass is the most abundant and renewable carbon source for many fungal species. The composition of biomass consists of about 40-45% cellulose, 20-30% hemicellulose, and 15-25% lignin and varies among plant species. In the bio-based industry, Aspergillus species and other fungi are used for the production of lignocellulolytic enzymes to pretreat agricultural waste biomass (e.g. wheat bran). In this study, we aimed to evaluate if it would be possible to create an Aspergillus strain that releases but does not metabolize hexoses from plant biomass. For this purpose, metabolic mutants were generated that were (partially) impaired in glycolysis, by deleting the hexokinase (hxkA) and glucokinase (glkA) genes. To prevent repression of enzyme production due to the accumulation of hexoses, strains were generated in which these mutations were combined with a mutation in creA, encoding the repressor involved in carbon catabolism. Phenotypic analysis revealed that growth of the ΔhxkAΔglkA mutant was reduced on wheat bran. However, hexoses did not accumulate during growth of the mutants on wheat bran, suggesting that glucose metabolism is re-routed towards alternative carbon catabolic pathways. Deletion of creA combined with blocking glycolysis resulted in an increased expression of pentose catabolic and phosphate pathway genes. This indicates that the reduced ability to use hexoses as carbon sources has resulted in a shift towards the pentose fraction of wheat bran as a major carbon source to support growth.
2018-02-26 | GSE94775 | GEO
Project description:Optimal glucose metabolism is central to the growth and development of cells. In microbial eukaryotes, carbon catabolite repression (CCR) mediates the preferential utilization of glucose, primarily by repressing alternate carbon source utilization. In fission yeast, CCR is mediated by transcriptional repressors Scr1 and the Tup/Ssn6 complex, with the Rst2 transcription factor important for activation of gluconeogenesis and sexual differentiation genes upon derepression. Through genetic and genome-wide methods, this study aimed to comprehensively characterize CCR in fission yeast by identifying the genes and biological processes that are regulated by Scr1, Tup/Ssn6 and Rst2, the core CCR machinery. The transcriptional response of S. pombe to glucose-sufficient or glucose-deficient growth conditions in wild type and CCR mutant cells was determined by RNA-seq and ChIP-seq. Scr1 was found to regulate genes involved in carbon metabolism, hexose uptake, gluconeogenesis and the TCA cycle. Surprisingly, a role for Scr1 in the suppression of sexual differentiation was also identified, as homothallic scr1 deletion mutants showed ectopic meiosis in carbon and nitrogen rich conditions. ChIP-seq characterised the targets of Tup/Ssn6 and Rst2 identifying regulatory roles within and independent of CCR. Finally, a subset of genes bound by all three factors was identified, implying that regulation of certain loci may be modulated in a competitive fashion between the Scr1, Tup/Ssn6 repressors and the Rst2 activator. By identifying the genes directly and indirectly regulated by Scr1, Tup/Ssn6 and Rst2, this study comprehensively defined the gene regulatory networks of CCR in fission yeast and revealed the transcriptional complexities which govern this system.
2018-12-12 | GSE123617 | GEO
Project description:Optimal glucose metabolism is central to the growth and development of cells. In microbial eukaryotes, carbon catabolite repression (CCR) mediates the preferential utilization of glucose, primarily by repressing alternate carbon source utilization. In fission yeast, CCR is mediated by transcriptional repressors Scr1 and the Tup/Ssn6 complex, with the Rst2 transcription factor important for activation of gluconeogenesis and sexual differentiation genes upon derepression. Through genetic and genome-wide methods, this study aimed to comprehensively characterize CCR in fission yeast by identifying the genes and biological processes that are regulated by Scr1, Tup/Ssn6 and Rst2, the core CCR machinery. The transcriptional response of S. pombe to glucose-sufficient or glucose-deficient growth conditions in wild type and CCR mutant cells was determined by RNA-seq and ChIP-seq. Scr1 was found to regulate genes involved in carbon metabolism, hexose uptake, gluconeogenesis and the TCA cycle. Surprisingly, a role for Scr1 in the suppression of sexual differentiation was also identified, as homothallic scr1 deletion mutants showed ectopic meiosis in carbon and nitrogen rich conditions. ChIP-seq characterised the targets of Tup/Ssn6 and Rst2 identifying regulatory roles within and independent of CCR. Finally, a subset of genes bound by all three factors was identified, implying that regulation of certain loci may be modulated in a competitive fashion between the Scr1, Tup/Ssn6 repressors and the Rst2 activator. By identifying the genes directly and indirectly regulated by Scr1, Tup/Ssn6 and Rst2, this study comprehensively defined the gene regulatory networks of CCR in fission yeast and revealed the transcriptional complexities which govern this system.
2018-12-12 | GSE123614 | GEO
Project description:Cre1 is an important transcription factor that regulates carbon catabolite repression (CCR) and is widely conserved across fungi. This gene has been extensively studied in several Ascomycota species, whereas its role in gene expression regulation in the Basidiomycota remains poorly understood. Here, we identified and investigated the role of cre1 in Coprinopsis cinerea, a basidiomycete model mushroom that can efficiently degrade lignocellulosic plant wastes. We used a rapid and efficient gene deletion approach based on PCR-amplified split-marker DNA cassettes together with in-vitro assembled Cas9-guide RNA ribonucleoproteins (Cas9-RNPs) to generate C. cinerea cre1 gene deletion strains. Gene expression profiling of two independent C. cinerea cre1 mutants showed significant deregulation of carbohydrate metabolism, plant cell wall degrading enzymes (PCWDEs), plasma membrane transporter-related and several transcription factor encoding genes, among others. Our results support the notion that, similarly to reports in the ascomycetes, Cre1 of C. cinerea orchestrates CCR through a combined regulation of diverse genes, including PCWDEs, transcription factors that positively regulate PCWDEs and membrane transporters which could import simple sugars that can induce the expression of PWCDEs. Somewhat paradoxically, though in accordance with other Agaricomycetes, genes related to lignin degradation were mostly downregulated in cre1 mutants, indicating they fall under different regulation than other PCWDEs. The gene deletion approach and the data presented in this paper expand our knowledge of CCR in the Basidiomycota and provide functional hypotheses on genes related to plant biomass degradation.
2022-11-01 | GSE205749 | GEO
Project description:The glucose dual-affinity transport system (low- and high-affinity) is a conserved strategy exerted by microorganisms to cope with the naturally fluctuating availability of nutrients in the environment. The glucose sensing and uptaking process were believed to be tightly involved in cellulases expression regulation in cellulolytic fungi. However, both the identities and functions of the major molecular components of this evolutionarily conserved system in filamentous fungi remain elusive. Here, we conducted a systematic identification and characterization of the glucose dual-affinity transport system in the model fungus Neurospora crassa. Using RNA sequencing coupled with functional transport analyses, we were able to assign GLT-1 (Km = 18.42 ± 3.38 mM) and HGT-1/-2 (Km = 16.13 ± 0.95 µM and 98.97 ± 22.02 µM) to low- and high-affinity glucose transport systems, respectively. The high-affinity transporters hgt-1/-2 were able to complement a moderate growth defect under high glucose when glt-1 was deleted. Simultaneous deletion of hgt-1/-2 led to extensive derepression of genes for plant cell wall deconstruction on cellulose. This suppression by HGT-1/-2 was connected to both carbon catabolite repression (CCR) and the cyclic adenosine monophosphate-protein kinase A pathway. Alteration of a residue conserved across taxa for hexose-transporters was found to result in a loss of glucose-transporting function, whereas CCR signal transduction was retained, indicating a dual function for HGT-1/-2 as “transceptors”. In this study, GLT-1 and HGT-1/-2 are identified as the key components of the glucose dual-affinity transport system, which play diverse roles in glucose transport and carbon metabolism. Given their wide conservation across fungal species, the glucose dual-affinity transport components and their pleiotropic roles revealed in this study would shed extensive new light on the molecular basis of nutrient transport, signaling, and plant cell wall degradation in fungi.
2017-01-04 | GSE78952 | GEO
Project description:The goals of this study are to compare differential gene expressions for Penicillium oxalicum wild type strain (WT), laeA knockout strain (ΔlaeA), creA knockout strain (ΔcreA), and double genes knockout strain (ΔlaeAΔcreA). The deletion of laeA downregulated genes involved in oxidation- reduction process, alkaloid metabolic process, and transmembrane transport. We find the expression levels of seven secondary metabolism gene clusters (totally 28 clusters) were silenced inΔlaeA. The deletion of creA upregulated genes involved in hydrolase activity, acting on glycosyl bonds. Many genes involved in conidiation were drastically regulated inΔlaeAΔcreA. This study provides the information that combined laeA and creA function are required in conidiation and hydrolase activity of P. oxalicum.
2014-11-19 | GSE63408 | GEO
Project description:In filamentous ascomycete fungi, the utilization of alternate carbon sources is influenced by the zinc finger transcription factor CreA/CRE-1, which encodes a carbon catabolite repressor protein homologous to Mig1 from Saccharomyces cerevisiae. In Neurospora crassa, deletion of cre-1 results in increased secretion of amylase and β-galactosidase. Here, we determined the CRE-1 regulon by investigating the transcriptome of a Δcre-1 strain compared to wild type when grown on Avicel versus minimal medium (MM). Our data provide comprehensive information on the CRE-1 regulon in N. crassa and contribute to deciphering the global role of carbon catabolite repression in filamentous ascomycete fungi during plant cell wall deconstruction.
2011-07-21 | GSE30313 | GEO