scholarly journals Xylanase Gene Transcription in Trichoderma reesei Is Triggered by Different Inducers Representing Different Hemicellulosic Pentose Polymers

2013 ◽  
Vol 12 (3) ◽  
pp. 390-398 ◽  
Author(s):  
Silvia Herold ◽  
Robert Bischof ◽  
Benjamin Metz ◽  
Bernhard Seiboth ◽  
Christian P. Kubicek
Keyword(s):  
Trichoderma Reesei ◽  
Plant Cell Wall ◽  
Glycosyl Hydrolase ◽  
Xylose Reductase ◽  
Genome Mining ◽  
Transcriptional Analysis ◽  
Cell Wall Degrading Enzymes ◽  
Content Type ◽  
Knockout Mutants ◽  
Catabolite Repressor

ABSTRACTThe ascomyceteTrichoderma reeseiis a paradigm for the regulation and production of plant cell wall-degrading enzymes, including xylanases. Four xylanases, including XYN1 and XYN2 of glycosyl hydrolase family 11 (GH11), the GH10 XYN3, and the GH30 XYN4, were already described. By genome mining, we identified a fifth xylanase, XYN5, belonging to GH11. Transcriptional analysis reveals that the expression of all xylanases butxyn3is induced byd-xylose, dependent on the cellulase and xylanase regulator XYR1 and negatively regulated by the carbon catabolite repressor CRE1. Impairment ofd-xylose catabolism at thed-xylose reductase and xylitol dehydrogenase step strongly enhanced induction byd-xylose. Knockout of thel-xylulose reductase-encoding genelxr3, which connects thed-xylose andl-arabinose catabolic pathways, had no effect on xylanase induction. Besides the induction byd-xylose, theT. reeseixylanases were also induced byl-arabinose, and this induction was also enhanced in knockout mutants inl-arabinose reductase (xyl1),l-arabitol dehydrogenase (lad1), andl-xylulose reductase (lxr3). Induction byl-arabinose was also XYR1 dependent. Analysis of intracellular polyols revealed accumulation of xylitol in all strains only during incubation withd-xylose and accumulation ofl-arabitol only during incubation withl-arabinose. Induction byl-arabinose could be further stimulated by addition ofd-xylose. We conclude that the expression of theT. reeseixylanases can be induced by bothd-xylose andl-arabinose, but independently of each other and by using different inducing metabolites.

scholarly journals Omics Analyses of Trichoderma reesei CBS999.97 and QM6a Indicate the Relevance of Female Fertility to Carbohydrate-Active Enzyme and Transporter Levels

2017 ◽  
Vol 83 (22) ◽  
Author(s):  
Doris Tisch ◽  
Kyle R. Pomraning ◽  
James R. Collett ◽  
Michael Freitag ◽  
Scott E. Baker ◽  
...  
Keyword(s):  
Cell Wall ◽  
Gene Regulation ◽  
Carbon Source ◽  
Trichoderma Reesei ◽  
Plant Cell Wall ◽  
Female Fertility ◽  
Mating Types ◽  
Carbon Source Utilization ◽  
Cell Wall Degrading Enzymes ◽  
Content Type

ABSTRACT The filamentous fungus Trichoderma reesei is found predominantly in the tropics but also in more temperate regions, such as Europe, and is widely known as a producer of large amounts of plant cell wall-degrading enzymes. We sequenced the genome of the sexually competent isolate CBS999.97, which is phenotypically different from the female sterile strain QM6a but can cross sexually with QM6a. Transcriptome data for growth on cellulose showed that entire carbohydrate-active enzyme (CAZyme) families are consistently differentially regulated between these strains. We evaluated backcrossed strains of both mating types, which acquired female fertility from CBS999.97 but maintained a mostly QM6a genetic background, and we could thereby distinguish between the effects of strain background and female fertility or mating type. We found clear regulatory differences associated with female fertility and female sterility, including regulation of CAZyme and transporter genes. Analysis of carbon source utilization, transcriptomes, and secondary metabolites in these strains revealed that only a few changes in gene regulation are consistently correlated with different mating types. Different strain backgrounds (QM6a versus CBS999.97) resulted in the most significant alterations in the transcriptomes and in carbon source utilization, with decreased growth of CBS999.97 on several amino acids (for example proline or alanine), which further correlated with the downregulation of genes involved in the respective pathways. In combination, our findings support a role of fertility-associated processes in physiology and gene regulation and are of high relevance for the use of sexual crossing in combining the characteristics of two compatible strains or quantitative trait locus (QTL) analysis. IMPORTANCE Trichoderma reesei is a filamentous fungus with a high potential for secretion of plant cell wall-degrading enzymes. We sequenced the genome of the fully fertile field isolate CBS999.97 and analyzed its gene regulation characteristics in comparison with the commonly used laboratory wild-type strain QM6a, which is not female fertile. Additionally, we also evaluated fully fertile strains with genotypes very close to that of QM6a in order to distinguish between strain-specific and fertility-specific characteristics. We found that QM6a and CBS999.97 clearly differ in their growth patterns on different carbon sources, CAZyme gene regulation, and secondary metabolism. Importantly, we found altered regulation of 90 genes associated with female fertility, including CAZyme genes and transporter genes, but only minor mating type-dependent differences. Hence, when using sexual crossing in research and for strain improvement, it is important to consider female fertile and female sterile strains for comparison with QM6a and to achieve optimal performance.

scholarly journals Draft Genome Sequence and Transcriptional Analysis of Rosellinia necatrix Infected with a Virulent Mycovirus

2018 ◽  
Vol 108 (10) ◽  
pp. 1206-1211 ◽  
Author(s):  
Takeo Shimizu ◽  
Satoko Kanematsu ◽  
Hajime Yaegashi
Keyword(s):  
Genome Sequence ◽  
Molecular Mechanisms ◽  
Plant Cell Wall ◽  
Draft Genome ◽  
Transcriptional Analysis ◽  
Draft Genome Sequence ◽  
Effective Control ◽  
Economic Losses ◽  
Rosellinia Necatrix ◽  
Cell Wall Degrading Enzymes

Understanding the molecular mechanisms of pathogenesis is useful in developing effective control methods for fungal diseases. The white root rot fungus Rosellinia necatrix is a soilborne pathogen that causes serious economic losses in various crops, including fruit trees, worldwide. Here, using next-generation sequencing techniques, we first produced a 44-Mb draft genome sequence of R. necatrix strain W97, an isolate from Japan, in which 12,444 protein-coding genes were predicted. To survey differentially expressed genes (DEGs) associated with the pathogenesis of the fungus, the hypovirulent W97 strain infected with Rosellinia necatrix megabirnavirus 1 (RnMBV1) was used for a comprehensive transcriptome analysis. In total, 545 and 615 genes are up- and down-regulated, respectively, in R. necatrix infected with RnMBV1. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses of the DEGs suggested that primary and secondary metabolism would be greatly disturbed in R. necatrix infected with RnMBV1. The genes encoding transcriptional regulators, plant cell wall-degrading enzymes, and toxin production, such as cytochalasin E, were also found in the DEGs. The genetic resources provided in this study will accelerate the discovery of genes associated with pathogenesis and other biological characteristics of R. necatrix, thus contributing to disease control.

scholarly journals A Novel Cys2His2 Zinc Finger Homolog of AZF1 Modulates Holocellulase Expression in Trichoderma reesei

mSystems ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Amanda Cristina Campos Antonieto ◽  
Karoline Maria Vieira Nogueira ◽  
Renato Graciano de Paula ◽  
Luísa Czamanski Nora ◽  
Murilo Henrique Anzolini Cassiano ◽  
...  
Keyword(s):  
Trichoderma Reesei ◽  
Filamentous Fungi ◽  
Plant Cell Wall ◽  
Plant Biomass ◽  
Regulatory Elements ◽  
Biomass Degradation ◽  
Control Of Gene Expression ◽  
Content Type ◽  
Encoding Genes

ABSTRACT Filamentous fungi are remarkable producers of enzymes dedicated to the degradation of sugar polymers found in the plant cell wall. Here, we integrated transcriptomic data to identify novel transcription factors (TFs) related to the control of gene expression of lignocellulosic hydrolases in Trichoderma reesei and Aspergillus nidulans. Using various sets of differentially expressed genes, we identified some putative cis-regulatory elements that were related to known binding sites for Saccharomyces cerevisiae TFs. Comparative genomics allowed the identification of six transcriptional factors in filamentous fungi that have corresponding S. cerevisiae homologs. Additionally, a knockout strain of T. reesei lacking one of these TFs (S. cerevisiae AZF1 homolog) displayed strong reductions in the levels of expression of several cellulase-encoding genes in response to both Avicel and sugarcane bagasse, revealing a new player in the complex regulatory network operating in filamentous fungi during plant biomass degradation. Finally, RNA sequencing (RNA-seq) analysis showed the scope of the AZF1 homologue in regulating a number of processes in T. reesei, and chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) provided evidence for the direct interaction of this TF in the promoter regions of cel7a, cel45a, and swo. Therefore, we identified here a novel TF which plays a positive effect in the expression of cellulase-encoding genes in T. reesei. IMPORTANCE In this work, we used a systems biology approach to map new regulatory interactions in Trichoderma reesei controlling the expression of genes encoding cellulase and hemicellulase. By integrating transcriptomics related to complex biomass degradation, we were able to identify a novel transcriptional regulator which is able to activate the expression of these genes in response to two different cellulose sources. In vivo experimental validation confirmed the role of this new regulator in several other processes related to carbon source utilization and nutrient transport. Therefore, this work revealed novel forms of regulatory interaction in this model system for plant biomass deconstruction and also represented a new approach that could be easy applied to other organisms.

scholarly journals d-Xylose as a Repressor or Inducer of Xylanase Expression in Hypocrea jecorina (Trichoderma reesei)

2010 ◽  
Vol 76 (6) ◽  
pp. 1770-1776 ◽  
Author(s):  
Astrid R. Mach-Aigner ◽  
Marion E. Pucher ◽  
Robert L. Mach
Keyword(s):  
Trichoderma Reesei ◽  
Xylose Reductase ◽  
Negative Influence ◽  
Hypocrea Jecorina ◽  
Xylanase Gene ◽  
Catabolite Repressor ◽  
Encoding Gene ◽  
Encoding Genes ◽  
Xylose Concentration

ABSTRACT For Hypocrea jecorina (anamorph Trichoderma reesei), a filamentous fungus used for hydrolase production in different industries, it has been a long-term practice to use d-xylose as an inducing substance. We demonstrate in this study that the degree of xylanase-encoding gene induction strictly depends on the concentration of d-xylose, which was found to be optimal from 0.5 to 1 mM for 3 h of cultivation. At higher concentrations of d-xylose, a reduced level of xylanase gene expression was observed. In the present study, we also provide evidence that the d-xylose concentration-dependent induction is antagonized by carbon catabolite repressor 1. This repressor mediates its influence on d-xylose indirectly, by reducing the expression of xylanase regulator 1, the main activator of most hydrolase-encoding genes. Additionally, a direct influence of the repressor on xylanase 1 expression in the presence of d-xylose was found. Furthermore, we show that d-xylose reductase 1 is needed to metabolize d-xylose to achieve full induction of xylanase expression. Finally, a strain which expresses xylanase regulator 1 at a constant level was used to partially overcome the negative influence exerted by carbon catabolite repressor 1 on d-xylose.

scholarly journals Discovery of 16-Demethylrifamycins by Removing the Predominant Polyketide Biosynthesis Pathway in Micromonospora sp. Strain TP-A0468

2018 ◽  
Vol 85 (4) ◽  
Author(s):  
Qiang Zhou ◽  
Guang-Cai Luo ◽  
Huizhan Zhang ◽  
Gong-Li Tang
Keyword(s):  
Natural Products ◽  
Transcriptional Activation ◽  
Metabolic Flux ◽  
Genome Mining ◽  
Gene Clusters ◽  
Bioinformatic Analysis ◽  
Transcriptional Analysis ◽  
Content Type ◽  
In The Wild ◽  
New Compounds

ABSTRACT A number of strategies have been developed to mine novel natural products based on biosynthetic gene clusters and there have been dozens of successful cases facilitated by the development of genomic sequencing. During our study on biosynthesis of the antitumor polyketide kosinostatin (KST), we found that the genome of Micromonospora sp. strain TP-A0468, the producer of KST, contains other potential polyketide gene clusters, with no encoded products detected. Deletion of kst cluster led to abolishment of KST and the enrichment of several new compounds, which were isolated and characterized as 16-demethylrifamycins (referred to here as compounds 3 to 6). Transcriptional analysis demonstrated that the expression of the essential genes related to the biosynthesis of compounds 3 to 6 was comparable to the level in the wild-type and in the kst cluster deletion strain. This indicates that the accumulation of these compounds was due to the redirection of metabolic flux rather than transcriptional activation. Genetic disruption, chemical complementation, and bioinformatic analysis revealed that the production of compounds 3 to 6 was accomplished by cross talk between the two distantly placed polyketide gene clusters pks3 and M-rif. This finding not only enriches the analogue pool and the biosynthetic diversity of rifamycins but also provides an auxiliary strategy for natural product discovery through genome mining in polyketide-producing microorganisms. IMPORTANCE Natural products are essential in the development of novel clinically used drugs. Discovering new natural products and modifying known compounds are still the two main ways to generate new candidates. Here, we have discovered several rifamycins with varied skeleton structures by redirecting the metabolic flux from the predominant polyketide biosynthetic pathway to the rifamycin pathway in the marine actinomycetes species Micromonospora sp. strain TP-A0468. Rifamycins are indispensable chemotherapeutics in the treatment of various diseases such as tuberculosis, leprosy, and AIDS-related mycobacterial infections. This study exemplifies a useful method for the discovery of cryptic natural products in genome-sequenced microbes. Moreover, the 16-demethylrifamycins and their genetically manipulable producer provide a new opportunity in the construction of novel rifamycin derivates to aid in the defense against the ever-growing drug resistance of Mycobacterium tuberculosis.

scholarly journals The Trk Potassium Transporter Is Required for RsmB-Mediated Activation of Virulence in the Phytopathogen Pectobacterium wasabiae

2015 ◽  
Vol 198 (2) ◽  
pp. 248-255 ◽  
Author(s):  
Rita S. Valente ◽  
Karina B. Xavier
Keyword(s):  
Cell Wall ◽  
Environmental Factor ◽  
Plant Cell ◽  
Plant Pathogen ◽  
Plant Cell Wall ◽  
Cell Wall Degrading Enzymes ◽  
Content Type ◽  
Regulatory Pathways ◽  
Potassium Transporter ◽  
Degrading Enzymes

ABSTRACTPectobacterium wasabiae(previously known asErwinia carotovora) is an important plant pathogen that regulates the production of plant cell wall-degrading enzymes through anN-acyl homoserine lactone-based quorum sensing system and through the GacS/GacA two-component system (also known as ExpS/ExpA). At high cell density, activation of GacS/GacA induces the expression of RsmB, a noncoding RNA that is essential for the activation of virulence in this bacterium. A genetic screen to identify regulators of RsmB revealed that mutants defective in components of a putative Trk potassium transporter (trkHandtrkA) had decreasedrsmBexpression. Further analysis of these mutants showed that changes in potassium concentration influencedrsmBexpression and consequent tissue damage in potato tubers and that this regulation required an intact Trk system. Regulation ofrsmBexpression by potassium via the Trk system occurred even in the absence of the GacS/GacA system, demonstrating that these systems act independently and are both required for full activation of RsmB and for the downstream induction of virulence in potato infection assays. Overall, our results identified potassium as an essential environmental factor regulating the Rsm system, and the consequent induction of virulence, in the plant pathogenP. wasabiae.IMPORTANCECrop losses from bacterial diseases caused by pectolytic bacteria are a major problem in agriculture. By studying the regulatory pathways involved in controlling the expression of plant cell wall-degrading enzymes inPectobacterium wasabiae, we showed that the Trk potassium transport system plays an important role in the regulation of these pathways. The data presented further identify potassium as an important environmental factor in the regulation of virulence in this plant pathogen. We showed that a reduction in virulence can be achieved by increasing the extracellular concentration of potassium. Therefore, this work highlights how elucidation of the mechanisms involved in regulating virulence can lead to the identification of environmental factors that can influence the outcome of infection.

scholarly journals Trichoderma reesei Isolated From Austrian Soil With High Potential for Biotechnological Application

2021 ◽  
Vol 12 ◽  
Author(s):  
Wolfgang Hinterdobler ◽  
Guofen Li ◽  
Katharina Spiegel ◽  
Samira Basyouni-Khamis ◽  
Markus Gorfer ◽  
...  
Keyword(s):  
Trichoderma Reesei ◽  
Plant Cell Wall ◽  
Plant Biomass ◽  
Strain Improvement ◽  
High Potential ◽  
Heterologous Proteins ◽  
Cell Wall Degrading Enzymes ◽  
Type Locus ◽  
Tropical Regions ◽  
Synonymous Snps

Fungi of the genus Trichoderma are of high importance for biotechnological applications, in biocontrol and for production of homologous and heterologous proteins. However, sexual crossing under laboratory conditions has so far only been achieved with the species Trichoderma reesei, which was so far only isolated from tropical regions. Our isolation efforts aimed at the collection of Trichoderma strains from Austrian soils surprisingly also yielded 12 strains of the species T. reesei, which was previously not known to occur in Europe. Their identity was confirmed with tef1- and rpb2-sequencing and phylogenetic analysis. They could clearly be distinguished from tropical strains including the common laboratory wildtypes by UP-PCR and genetic variations adjacent to the mating type locus. The strains readily mated with reference strains derived from CBS999.97. Secreted cellulase and xylanase levels of these isolates were up to six-fold higher than those of QM6a indicating a high potential for strain improvement. The strains showed different responses to injury in terms of induction of sporulation, but a correlation to alterations in the nox1-gene sequence was not detected. Several synonymous SNPs were found in the sequence of the regulator gene noxR of the soil isolates compared to QM6a. Only in one strain, non-synonymous SNPs were found which impact a PEST sequence of NoxR, suggesting altered protein stability. The availability of sexually fertile strains from middle Europe naturally producing decent amounts of plant cell wall degrading enzymes opens up novel perspectives for non-GMO strain improvement and biological pretreatment of plant biomass for bioethanol production. Moreover, the varied response of these strains to injury in terms of sporulation, which is independent of Nox1 and NoxR suggests that additional regulators impact this phenomenon in T. reesei.

scholarly journals Erwinia carotovora Quorum Sensing System Regulates Host-Specific Virulence Factors and Development Delay in Drosophila melanogaster

mBio ◽  
2020 ◽  
Vol 11 (3) ◽  
Author(s):  
Filipe J. D. Vieira ◽  
Pol Nadal-Jimenez ◽  
Luis Teixeira ◽  
Karina B. Xavier
Keyword(s):  
Cell Wall ◽  
Drosophila Melanogaster ◽  
Quorum Sensing ◽  
Virulence Factors ◽  
Plant Cell Wall ◽  
Erwinia Carotovora ◽  
Homoserine Lactone ◽  
Cell Wall Degrading Enzymes ◽  
Content Type ◽  
Pectolytic Enzymes

ABSTRACT Multihost bacteria have to rapidly adapt to drastic environmental changes, relying on a fine integration of multiple stimuli for an optimal genetic response. Erwinia carotovora spp. are phytopathogens that cause soft-rot disease. Strain Ecc15 in particular is a model for bacterial oral-route infection in Drosophila melanogaster as it harbors a unique gene, evf, that encodes the Erwinia virulence factor (Evf), which is a major determinant for infection of the D. melanogaster gut. However, the factors involved in the regulation of evf expression are poorly understood. We investigated whether evf could be controlled by quorum sensing as, in the Erwinia genus, quorum sensing regulates pectolytic enzymes, the major virulence factors needed to infect plants. Here, we show that transcription of evf is positively regulated by quorum sensing in Ecc15 via acyl-homoserine lactone (AHL) signal synthase ExpI and AHL receptors ExpR1 and ExpR2. We also show that the load of Ecc15 in the gut depends upon the quorum sensing-mediated regulation of evf. Furthermore, we demonstrate that larvae infected with Ecc15 suffer a developmental delay as a direct consequence of the regulation of evf via quorum sensing. Finally, we demonstrate that evf is coexpressed with plant cell wall-degrading enzymes (PCWDE) during plant infection in a quorum sensing-dependent manner. Overall, our results show that Ecc15 relies on quorum sensing to control production of both pectolytic enzymes and Evf. This regulation influences the interaction of Ecc15 with its two known hosts, indicating that quorum sensing signaling may impact bacterial dissemination via insect vectors that feed on rotting plants. IMPORTANCE Integration of genetic networks allows bacteria to rapidly adapt to changing environments. This is particularly important in bacteria that interact with multiple hosts. Erwinia carotovora is a plant pathogen that uses Drosophila melanogaster as a vector. To interact with these two hosts, Ecc15 uses different sets of virulence factors: plant cell wall-degrading enzymes to infect plants and the Erwinia virulence factor (evf) to infect Drosophila. Our work shows that, despite the virulence factors being specific for each host, both sets are coactivated by homoserine lactone quorum sensing and by the two-component GacS/A system in infected plants. This regulation is essential for Ecc15 loads in the gut of Drosophila and minimizes the developmental delay caused by the bacteria with respect to the insect vector. Our findings provide evidence that coactivation of the host-specific factors in the plant may function as a predictive mechanism to maximize the probability of transit of the bacteria between hosts.

scholarly journals Coordinated and Distinct Functions of Velvet Proteins in Fusarium verticillioides

2014 ◽  
Vol 13 (7) ◽  
pp. 909-918 ◽  
Author(s):  
Nan Lan ◽  
Hanxing Zhang ◽  
Chengcheng Hu ◽  
Wenzhao Wang ◽  
Ana M. Calvo ◽  
...  
Keyword(s):  
Gene Knockout ◽  
Affinity Purification ◽  
Fusarium Verticillioides ◽  
Toxin Production ◽  
Transcriptional Analysis ◽  
Phenotypic Characterization ◽  
Morphological Development ◽  
Content Type ◽  
Knockout Mutants ◽  
Fungal Kingdom

ABSTRACTVelvet-domain-containing proteins are broadly distributed within the fungal kingdom. In the corn pathogenFusarium verticillioides, previous studies showed that the velvet proteinF. verticillioidesVE1 (FvVE1) is critical for morphological development, colony hydrophobicity, toxin production, and pathogenicity. In this study, tandem affinity purification of FvVE1 revealed that FvVE1 can form a complex with the velvet proteinsF. verticillioidesVelB (FvVelB) and FvVelC. Phenotypic characterization of gene knockout mutants showed that, as in the case of FvVE1, FvVelB regulated conidial size, hyphal hydrophobicity, fumonisin production, and oxidant resistance, while FvVelC was dispensable for these biological processes. Comparative transcriptional analysis of eight genes involved in the ROS (reactive oxygen species) removal system revealed that both FvVE1 and FvVelB positively regulated the transcription of a catalase-encoding gene,F. verticillioidesCAT2(FvCAT2). Deletion ofFvCAT2resulted in reduced oxidant resistance, providing further explanation of the regulation of oxidant resistance by velvet proteins in the fungal kingdom.

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