ABSTRACT
Fungi of the order Pucciniales are obligate plant biotrophs causing rust diseases. They exhibit a complex life cycle with the production of up to five spore types, infection of two unrelated hosts and an overwintering stage. Transcription factors (TFs) are key regulators of gene expression in eukaryote cells. In order to better understand genetic programs expressed during major transitions of the rust life cycle, we surveyed the complement of TFs in fungal genomes with an emphasis on Pucciniales. We found that despite their large gene numbers, rust genomes have a reduced repertoire of TFs compared to other fungi. The proportions of C2H2 and Zinc cluster - two of the most represented TF families in fungi - indicate differences in their evolutionary relationships in Pucciniales and other fungal taxa. The regulatory gene family encoding cold shock protein (CSP) showed a striking expansion in Pucciniomycotina with specific duplications in the order Pucciniales. The survey of expression profiles collected by transcriptomics along the life cycle of the poplar rust fungus revealed TF genes related to major biological transitions, e.g. response to environmental cues and host infection. Particularly, poplar rust CSPs were strongly expressed in basidia produced after the overwintering stage suggesting a possible role in dormancy exit. Expression during transition from dormant telia to basidia confirmed the specific expression of the three poplar rust CSP genes. Their heterologous expression in yeast improved cell growth after cold stress exposure, suggesting a probable regulatory function when the poplar rust fungus exits dormancy. This study addresses for the first time TF and regulatory genes involved in developmental transition in the rust life cycle opening perspectives to further explore molecular regulation in the biology of the Pucciniales.
Subject(s)
Basidiomycota , Populus , Animals , Basidiomycota/genetics , Cold Shock Proteins and Peptides/genetics , Cold Shock Proteins and Peptides/metabolism , Life Cycle Stages , Plant Diseases/microbiology , Populus/microbiology , Transcription Factors/genetics , Transcription Factors/metabolism , Transcriptome/geneticsABSTRACT
Many antibiotics, chemotherapeutics, crop protection agents and food preservatives originate from molecules produced by bacteria, fungi or plants. In recent years, genome mining methodologies have been widely adopted to identify and characterize the biosynthetic gene clusters encoding the production of such compounds. Since 2011, the 'antibiotics and secondary metabolite analysis shell-antiSMASH' has assisted researchers in efficiently performing this, both as a web server and a standalone tool. Here, we present the thoroughly updated antiSMASH version 4, which adds several novel features, including prediction of gene cluster boundaries using the ClusterFinder method or the newly integrated CASSIS algorithm, improved substrate specificity prediction for non-ribosomal peptide synthetase adenylation domains based on the new SANDPUMA algorithm, improved predictions for terpene and ribosomally synthesized and post-translationally modified peptides cluster products, reporting of sequence similarity to proteins encoded in experimentally characterized gene clusters on a per-protein basis and a domain-level alignment tool for comparative analysis of trans-AT polyketide synthase assembly line architectures. Additionally, several usability features have been updated and improved. Together, these improvements make antiSMASH up-to-date with the latest developments in natural product research and will further facilitate computational genome mining for the discovery of novel bioactive molecules.
Subject(s)
Secondary Metabolism/genetics , Software , Algorithms , Anti-Bacterial Agents/biosynthesis , Biological Products/metabolism , Biosynthetic Pathways/genetics , Codon , Genes , Internet , Peptide Synthases/metabolism , Peptides/chemistry , Peptides/metabolism , Polyketide Synthases/chemistry , Protein Domains , Protein Processing, Post-Translational , Terpenes/chemistryABSTRACT
[This corrects the article DOI: 10.1371/journal.pgen.1004496.].
ABSTRACT
Production of basidiomycete atromentin-derived pigments like variegatic acid (pulvinic acid-type) and involutin (diarylcyclopentenone) from the brown-rotter Serpula lacrymans and the ectomycorrhiza-forming Paxillus involutus, respectively, is induced by complex nutrition, and in the case of S. lacrymans, bacteria. Pigmentation in S. lacrymans was stimulated by 13 different bacteria and cell-wall-damaging enzymes (lytic enzymes and proteases), but not by lysozyme or mechanical damage. The use of protease inhibitors with Bacillus subtilis or heat-killed bacteria during co-culturing with S. lacrymans significantly reduced pigmentation indicating that enzymatic hyphal damage and/or released peptides, rather than mechanical injury, was the major cause of systemic pigment induction. Conversely, no significant pigmentation by bacteria was observed from P. involutus. We found additional putative transcriptional composite elements of atromentin synthetase genes in P. involutus and other ectomycorrhiza-forming species that were absent from S. lacrymans and other brown-rotters. Variegatic and its precursor xerocomic acid, but not involutin, in return inhibited swarming and colony biofilm spreading of Bacillus subtilis, but did not kill B. subtilis. We suggest that dissimilar pigment regulation by fungal lifestyle was a consequence of pigment bioactivity and additional promoter motifs. The focus on basidiomycete natural product gene induction and regulation will assist in future studies to determine global regulators, signalling pathways and associated transcription factors of basidiomycetes.
Subject(s)
Agaricales/metabolism , Bacterial Physiological Phenomena , Biofilms/growth & development , Gene Expression Regulation, Fungal , Microbial Interactions/physiology , Pigments, Biological/genetics , Agaricales/classification , Agaricales/genetics , Agaricales/growth & development , Bacillus subtilis/growth & development , Bacillus subtilis/physiology , Bacteria/genetics , Bacteria/growth & development , Bacteria/metabolism , Benzoquinones/metabolism , Cell Wall/metabolism , Coculture Techniques , Computer Simulation , Conserved Sequence , Databases, Genetic , Fungal Proteins/genetics , Microbial Interactions/genetics , Multigene Family/genetics , Phenols/metabolism , Pigments, Biological/biosynthesis , Pigments, Biological/metabolism , Promoter Regions, GeneticABSTRACT
Melanins play a crucial role in defending organisms against external stressors. In several pathogenic fungi, including the human pathogen Aspergillus fumigatus, melanin production was shown to contribute to virulence. A. fumigatus produces two different types of melanins, i.e., pyomelanin and dihydroxynaphthalene (DHN)-melanin. DHN-melanin forms the gray-green pigment characteristic for conidia, playing an important role in immune evasion of conidia and thus for fungal virulence. The DHN-melanin biosynthesis pathway is encoded by six genes organized in a cluster with the polyketide synthase gene pksP as a core element. Here, cross-species promoter analysis identified specific DNA binding sites in the DHN-melanin biosynthesis genes pksP-arp1 intergenic region that can be recognized by bHLH and MADS-box transcriptional regulators. Independent deletion of two genes coding for the transcription factors DevR (bHLH) and RlmA (MADS-box) interfered with sporulation and reduced the expression of the DHN-melanin gene cluster. In vitro and in vivo experiments proved that these transcription factors cooperatively regulate pksP expression acting both as repressors and activators in a mutually exclusive manner. The dual role executed by each regulator depends on specific DNA motifs recognized in the pksP promoter region.
Subject(s)
Aspergillus fumigatus/metabolism , Basic Helix-Loop-Helix Transcription Factors/metabolism , Melanins/biosynthesis , Aspergillus fumigatus/genetics , Basic Helix-Loop-Helix Transcription Factors/genetics , Biosynthetic Pathways , Fungal Proteins/metabolism , Genes, Fungal , Melanins/genetics , Melanins/metabolism , Multigene Family , Pigmentation , Protein Binding , Protein Domains , Spores, Fungal/genetics , Spores, Fungal/metabolismABSTRACT
Aspergillus fumigatus is the predominant airborne pathogenic fungus causing invasive aspergillosis in immunocompromised patients. During infection A. fumigatus has to adapt to oxygen-limiting conditions in inflammatory or necrotic tissue. Previously, we identified a mitochondrial protein to be highly up-regulated during hypoxic adaptation. Here, this protein was found to represent the novel oxidoreductase HorA. In Saccharomyces cerevisiae a homologue was shown to play a role in biosynthesis of coenzyme Q. Consistently, reduced coenzyme Q content in the generated ΔhorA mutant indicated a respective function in A. fumigatus. Since coenzyme Q is involved in cellular respiration and maintaining cellular redox homeostasis, the strain ΔhorA displayed an impaired response to both oxidative and reductive stress, a delay in germination and an accumulation of NADH. Moreover, an increased resistance against antifungal drugs was observed. All phenotypes were completely reversed by the addition of the synthetic electron carrier menadione. The deletion strain ΔhorA showed significantly attenuated virulence in two murine infection models of invasive pulmonary aspergillosis. Therefore, the biosynthesis of coenzyme Q and, particularly, the fungal-specific protein HorA play a crucial role in virulence of A. fumigatus. Due to its absence in mammals, HorA might represent a novel therapeutic target against fungal infections.
Subject(s)
Aspergillus fumigatus/metabolism , Azoles/pharmacology , Oxidoreductases/metabolism , Ubiquinone/analogs & derivatives , Animals , Antifungal Agents/pharmacology , Aspergillus fumigatus/drug effects , Aspergillus fumigatus/enzymology , Aspergillus fumigatus/genetics , Cell Hypoxia/physiology , Disease Models, Animal , Female , Fungal Proteins/metabolism , Gene Deletion , Invasive Pulmonary Aspergillosis/microbiology , Mice , Oxidoreductases/genetics , Ubiquinone/biosynthesis , VirulenceABSTRACT
MOTIVATION: Secondary metabolites (SM) are structurally diverse natural products of high pharmaceutical importance. Genes involved in their biosynthesis are often organized in clusters, i.e., are co-localized and co-expressed. In silico cluster prediction in eukaryotic genomes remains problematic mainly due to the high variability of the clusters' content and lack of other distinguishing sequence features. RESULTS: We present Cluster Assignment by Islands of Sites (CASSIS), a method for SM cluster prediction in eukaryotic genomes, and Secondary Metabolites by InterProScan (SMIPS), a tool for genome-wide detection of SM key enzymes ('anchor' genes): polyketide synthases, non-ribosomal peptide synthetases and dimethylallyl tryptophan synthases. Unlike other tools based on protein similarity, CASSIS exploits the idea of co-regulation of the cluster genes, which assumes the existence of common regulatory patterns in the cluster promoters. The method searches for 'islands' of enriched cluster-specific motifs in the vicinity of anchor genes. It was validated in a series of cross-validation experiments and showed high sensitivity and specificity. AVAILABILITY AND IMPLEMENTATION: CASSIS and SMIPS are freely available at https://sbi.hki-jena.de/cassis CONTACT: thomas.wolf@leibniz-hki.de or ekaterina.shelest@leibniz-hki.de SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Subject(s)
Multigene Family , Peptide Synthases/genetics , Polyketide Synthases/genetics , Eukaryota , SoftwareABSTRACT
Lichtheimia species are the second most important cause of mucormycosis in Europe. To provide broader insights into the molecular basis of the pathogenicity-associated traits of the basal Mucorales, we report the full genome sequence of L. corymbifera and compared it to the genome of Rhizopus oryzae, the most common cause of mucormycosis worldwide. The genome assembly encompasses 33.6 MB and 12,379 protein-coding genes. This study reveals four major differences of the L. corymbifera genome to R. oryzae: (i) the presence of an highly elevated number of gene duplications which are unlike R. oryzae not due to whole genome duplication (WGD), (ii) despite the relatively high incidence of introns, alternative splicing (AS) is not frequently observed for the generation of paralogs and in response to stress, (iii) the content of repetitive elements is strikingly low (<5%), (iv) L. corymbifera is typically haploid. Novel virulence factors were identified which may be involved in the regulation of the adaptation to iron-limitation, e.g. LCor01340.1 encoding a putative siderophore transporter and LCor00410.1 involved in the siderophore metabolism. Genes encoding the transcription factors LCor08192.1 and LCor01236.1, which are similar to GATA type regulators and to calcineurin regulated CRZ1, respectively, indicating an involvement of the calcineurin pathway in the adaption to iron limitation. Genes encoding MADS-box transcription factors are elevated up to 11 copies compared to the 1-4 copies usually found in other fungi. More findings are: (i) lower content of tRNAs, but unique codons in L. corymbifera, (ii) Over 25% of the proteins are apparently specific for L. corymbifera. (iii) L. corymbifera contains only 2/3 of the proteases (known to be essential virulence factors) in comparison to R. oryzae. On the other hand, the number of secreted proteases, however, is roughly twice as high as in R. oryzae.
Subject(s)
Evolution, Molecular , Genome, Fungal , Mucorales/genetics , Mucormycosis/genetics , Alternative Splicing/genetics , Gene Duplication , Genomics , Humans , Mucorales/pathogenicity , Mucormycosis/microbiology , Virulence Factors/genetics , Virulence Factors/isolation & purificationABSTRACT
BACKGROUND: The Rhynchosporium species complex consists of hemibiotrophic fungal pathogens specialized to different sweet grass species including the cereal crops barley and rye. A sexual stage has not been described, but several lines of evidence suggest the occurrence of sexual reproduction. Therefore, a comparative genomics approach was carried out to disclose the evolutionary relationship of the species and to identify genes demonstrating the potential for a sexual cycle. Furthermore, due to the evolutionary very young age of the five species currently known, this genus appears to be well-suited to address the question at the molecular level of how pathogenic fungi adapt to their hosts. RESULTS: The genomes of the different Rhynchosporium species were sequenced, assembled and annotated using ab initio gene predictors trained on several fungal genomes as well as on Rhynchosporium expressed sequence tags. Structures of the rDNA regions and genome-wide single nucleotide polymorphisms provided a hypothesis for intra-genus evolution. Homology screening detected core meiotic genes along with most genes crucial for sexual recombination in ascomycete fungi. In addition, a large number of cell wall-degrading enzymes that is characteristic for hemibiotrophic and necrotrophic fungi infecting monocotyledonous hosts were found. Furthermore, the Rhynchosporium genomes carry a repertoire of genes coding for polyketide synthases and non-ribosomal peptide synthetases. Several of these genes are missing from the genome of the closest sequenced relative, the poplar pathogen Marssonina brunnea, and are possibly involved in adaptation to the grass hosts. Most importantly, six species-specific genes coding for protein effectors were identified in R. commune. Their deletion yielded mutants that grew more vigorously in planta than the wild type. CONCLUSION: Both cryptic sexuality and secondary metabolites may have contributed to host adaptation. Most importantly, however, the growth-retarding activity of the species-specific effectors suggests that host adaptation of R. commune aims at extending the biotrophic stage at the expense of the necrotrophic stage of pathogenesis. Like other apoplastic fungi Rhynchosporium colonizes the intercellular matrix of host leaves relatively slowly without causing symptoms, reminiscent of the development of endophytic fungi. Rhynchosporium may therefore become an object for studying the mutualism-parasitism transition.
Subject(s)
Ascomycota/classification , Ascomycota/genetics , Genome, Fungal , Genomics , Host Specificity , Phylogeny , Poaceae/microbiology , Amino Acid Sequence , Ascomycota/metabolism , DNA, Intergenic , Gene Expression Regulation, Fungal , Genes, Fungal , Genomics/methods , Multigene Family , Secondary Metabolism/geneticsABSTRACT
Basidiomycete fungi are characterized ecologically for their vital functional role in ecosystem carbon recycling and chemically for their capacity to produce a diverse array of small molecules. Chromophoric natural products derived from the quinone precursor atromentin, such as variegatic acid and involutin, have been shown to function in redox cycling. Yet, in the context of an inter-kingdom natural system these pigments are still elusive. Here, we co-cultured the model saprotrophic basidiomycete Serpula lacrymans with an ubiquitous terrestrial bacterium, either Bacillus subtilis, Pseudomonas putida, or Streptomyces iranensis. For each, there was induction of the gene cluster encoding a non-ribosomal peptide synthetase-like enzyme (atromentin synthetase) and an aminotransferase which together produce atromentin. Correspondingly, during co-culturing there was an increase in secreted atromentin-derived pigments, i.e., variegatic, xerocomic, isoxerocomic, and atromentic acid. Bioinformatic analyses from 14 quinone synthetase genes, twelve of which are encoded in a cluster, identified a common promoter motif indicating a general regulatory mechanism for numerous basidiomycetes.
Subject(s)
Bacillus subtilis/physiology , Basidiomycota/metabolism , Benzoquinones/metabolism , Pigments, Biological/biosynthesis , Pseudomonas putida/physiology , Streptomyces/physiology , Bacillus subtilis/growth & development , Basidiomycota/genetics , Basidiomycota/growth & development , Coculture Techniques , Multigene Family , Oxidation-Reduction , Pseudomonas putida/growth & development , Streptomyces/growth & developmentABSTRACT
BACKGROUND: Polyketide synthase (PKS) catalyzes the biosynthesis of polyketides, which are structurally and functionally diverse natural products in microorganisms and plants. Here, we have analyzed available full genome sequences of microscopic and macroscopic algae for the presence of type I PKS genes. RESULTS: Type I PKS genes are present in 15 of 32 analyzed algal species. In chlorophytes, large proteins in the MDa range are predicted in most sequenced species, and PKSs with free-standing acyltransferase domains (trans-AT PKSs) predominate. In a phylogenetic tree, PKS sequences from different algal phyla form clades that are distinct from PKSs from other organisms such as non-photosynthetic protists or cyanobacteria. However, intermixing is observed in some cases, for example polyunsaturated fatty acid (PUFA) and glycolipid synthases of various origins. Close relationships between type I PKS modules from different species or between modules within the same multimodular enzyme were identified, suggesting module duplications during evolution of algal PKSs. In contrast to type I PKSs, nonribosomal peptide synthetases (NRPSs) are relatively rare in algae (occurrence in 7 of 32 species). CONCLUSIONS: Our phylogenetic analysis of type I PKSs in algae supports an evolutionary scenario whereby integrated AT domains were displaced to yield trans-AT PKSs. Together with module duplications, the displacement of AT domains may constitute a major mechanism of PKS evolution in algae. This study advances our understanding of the diversity of eukaryotic PKSs and their evolutionary trajectories.
Subject(s)
Polyketide Synthases/metabolism , Chlorophyta/enzymology , Dinoflagellida/enzymology , Fatty Acids, Unsaturated/metabolism , Microalgae/enzymology , Polyketide Synthases/genetics , Substrate SpecificityABSTRACT
Fungi produce α-aminoadipate, a precursor for penicillin and lysine via the α-aminoadipate pathway. Despite the biotechnological importance of this pathway, the essential isomerization of homocitrate via homoaconitate to homoisocitrate has hardly been studied. Therefore, we analysed the role of homoaconitases and aconitases in this isomerization. Although we confirmed an essential contribution of homoaconitases from Saccharomyces cerevisiae and Aspergillus fumigatus, these enzymes only catalysed the interconversion between homoaconitate and homoisocitrate. In contrast, aconitases from fungi and the thermophilic bacterium Thermus thermophilus converted homocitrate to homoaconitate. Additionally, a single aconitase appears essential for energy metabolism, glutamate and lysine biosynthesis in respirating filamentous fungi, but not in the fermenting yeast S. cerevisiae that possesses two contributing aconitases. While yeast Aco1p is essential for the citric acid cycle and, thus, for glutamate synthesis, Aco2p specifically and exclusively contributes to lysine biosynthesis. In contrast, Aco2p homologues present in filamentous fungi were transcribed, but enzymatically inactive, revealed no altered phenotype when deleted and did not complement yeast aconitase mutants. From these results we conclude that the essential requirement of filamentous fungi for respiration versus the preference of yeasts for fermentation may have directed the evolution of aconitases contributing to energy metabolism and lysine biosynthesis.
Subject(s)
2-Aminoadipic Acid/metabolism , Aconitate Hydratase/metabolism , Fungi/enzymology , Fungi/metabolism , Lysine/biosynthesis , Tricarboxylic Acids/metabolism , Biosynthetic Pathways/genetics , Energy MetabolismABSTRACT
Fungal hybrid enzymes consisting of a polyketide synthase (PKS) and a nonribosomal peptide synthetase (NRPS) module are involved in the biosynthesis of a vast array of ecologically and medicinally relevant natural products. Whereas a dozen gene clusters could be assigned to the requisite PKS-NRPS pathways, the programming of the multifunctional enzymes is still enigmatic. Through engineering and heterologously expressing a chimera of PKS (lovastatin synthase, LovB) and NRPS (cytochalasin synthase, CheA) in Aspergillus terreus, we noted the potential incompatibility of a fungal highly reducing PKS (hrPKS) with the NRPS component of fungal PKS-NRPS hybrids. To rationalize the unexpected outcome of the gene fusion experiments, we conducted extensive bioinformatic analyses of fungal PKS-NRPS hybrids and LovB-type PKS. From motif studies and the function of the engineered chimeras, a noncanonical function of C-terminal condensation (C) domains in truncated PKS-NRPS homologues was inferred. More importantly, sequence alignments and phylogenetic trees revealed an evolutionary imprint of the PKS-NRPS domains, which reflect the evolutionary history of the entire megasynthase. Furthermore, a detailed investigation of C and adenylation (A) domains provides support for a scenario in which not only the A domain but also the C domain participates in amino acid selection. These findings shed new light on the complex code of this emerging class of multifunctional enzymes and will greatly facilitate future combinatorial biosynthesis and pathway engineering approaches towards natural product analogues.
Subject(s)
Aspergillus/enzymology , Biological Evolution , Peptide Synthases/metabolism , Polyketide Synthases/metabolism , Amino Acids/chemistry , Aspergillus/genetics , Aspergillus/metabolism , Biocatalysis , Computational Biology , Indole Alkaloids/chemistry , Lovastatin/chemistry , Models, Molecular , Molecular Conformation , Peptide Synthases/analysis , Peptide Synthases/genetics , Phylogeny , Polyketide Synthases/analysis , Polyketide Synthases/genetics , Polyketides/chemistry , Sequence AlignmentABSTRACT
MOTIVATION: Prediction of transcription factor binding sites (TFBSs) is crucial for promoter modeling and network inference. Quality of the predictions is spoiled by numerous false positives, which persist as the main problem for all presently available TFBS search methods. RESULTS: We suggest a novel approach, which is alternative to widely used position weight matrices (PWMs) and Hidden Markov Models. Each motif of the input set is used as a search template to scan a query sequence. Found motifs are assigned scores depending on the non-randomness of the motif's occurrence, the number of matching searching motifs and the number of mismatches. The non-randomness is estimated by comparison of observed numbers of matching motifs with those predicted to occur by chance. The latter can be calculated given the base compositions of the motif and the query sequence. The method does not require preliminary alignment of the input motifs, hence avoiding uncertainties introduced by the alignment procedure. In comparison with PWM-based tools, our method demonstrates higher precision by the same sensitivity and specificity. It also tends to outperform methods combining pattern and PWM search. Most important, it allows reducing the number of false positive predictions significantly. AVAILABILITY: The method is implemented in a tool called SiTaR (Site Tracking and Recognition) and is available at http://sbi.hki-jena.de/sitar/index.php. CONTACT: ekaterina.shelest@hki-jena.de SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Subject(s)
Promoter Regions, Genetic , Sequence Analysis, DNA , Software , Transcription Factors/metabolism , Binding Sites , Nucleotide Motifs , Sensitivity and SpecificityABSTRACT
The genomes of filamentous fungi comprise numerous putative gene clusters coding for the biosynthesis of chemically and structurally diverse secondary metabolites (SMs), which are rarely expressed under laboratory conditions. Previous approaches to activate these genes were based primarily on artificially targeting the cellular protein synthesis apparatus. Here, we applied an alternative approach of genetically impairing the protein degradation apparatus of the model fungus Aspergillus nidulans by deleting the conserved eukaryotic csnE/CSN5 deneddylase subunit of the COP9 signalosome. This defect in protein degradation results in the activation of a previously silenced gene cluster comprising a polyketide synthase gene producing the antibiotic 2,4-dihydroxy-3-methyl-6-(2-oxopropyl)benzaldehyde (DHMBA). The csnE/CSN5 gene is highly conserved in fungi, and therefore, the deletion is a feasible approach for the identification of new SMs.
Subject(s)
Aspergillus nidulans/genetics , Biological Products/metabolism , Gene Expression Regulation, Fungal , Gene Silencing , Multigene Family , Aspergillus nidulans/metabolism , COP9 Signalosome Complex , Gene Deletion , Metalloendopeptidases/genetics , Metalloendopeptidases/metabolism , Multiprotein Complexes/genetics , Multiprotein Complexes/metabolism , Peptide Hydrolases/genetics , Peptide Hydrolases/metabolismABSTRACT
Fungi produce numerous low molecular weight molecules endowed with a multitude of biological activities. However, mining the full-genome sequences of fungi indicates that their potential to produce secondary metabolites is greatly underestimated. Because most of the biosynthesis gene clusters are silent under laboratory conditions, one of the major challenges is to understand the physiological conditions under which these genes are activated. Thus, we cocultivated the important model fungus Aspergillus nidulans with a collection of 58 soil-dwelling actinomycetes. By microarray analyses of both Aspergillus secondary metabolism and full-genome arrays and Northern blot and quantitative RT-PCR analyses, we demonstrate at the molecular level that a distinct fungal-bacterial interaction leads to the specific activation of fungal secondary metabolism genes. Most surprisingly, dialysis experiments and electron microscopy indicated that an intimate physical interaction of the bacterial and fungal mycelia is required to elicit the specific response. Gene knockout experiments provided evidence that one induced gene cluster codes for the long-sought after polyketide synthase (PKS) required for the biosynthesis of the archetypal polyketide orsellinic acid, the typical lichen metabolite lecanoric acid, and the cathepsin K inhibitors F-9775A and F-9775B. A phylogenetic analysis demonstrates that orthologs of this PKS are widespread in nature in all major fungal groups, including mycobionts of lichens. These results provide evidence of specific interaction among microorganisms belonging to different domains and support the hypothesis that not only diffusible signals but intimate physical interactions contribute to the communication among microorganisms and induction of otherwise silent biosynthesis genes.
Subject(s)
Actinobacteria/physiology , Aspergillus nidulans/genetics , Aspergillus nidulans/metabolism , Macrolides/metabolism , Actinobacteria/classification , Actinobacteria/ultrastructure , Aspergillus nidulans/ultrastructure , Blotting, Northern , Chromatography, High Pressure Liquid , Ecosystem , Fungal Proteins/genetics , Fungal Proteins/metabolism , Gene Expression Profiling , Gene Expression Regulation, Fungal , Genome, Fungal/genetics , Microscopy, Electron, Scanning , Molecular Structure , Mutation , Mycelium/genetics , Mycelium/metabolism , Mycelium/ultrastructure , Oligonucleotide Array Sequence Analysis , Phylogeny , Polyketide Synthases/classification , Polyketide Synthases/genetics , Polyketide Synthases/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Salicylates/analysis , Salicylates/chemistry , Zearalenone/chemistry , Zearalenone/metabolismABSTRACT
SUMMARY: The state of the art in promoter modeling for higher eukaryotes is predicting not single transcription factor binding sites (TFBSs), but their combinations. The new tool utilizes a previously developed method of distance distributions of TFBS pairs. We model the random distribution of distances and compare it with the distribution observed in the query sequences. Comparison of the profiles allows filtering out the 'noise' and retaining the potentially functional combinations. This approach has proved its usefulness as a filtering technique for the selection of TFBS pairs for promoter modeling and is now implemented as a tool in R. As an input, it can use the outputs of three different TFBS- and motif-predictive tools (Gibbs Sampler for motifs, Match and MEME/FIMO for PWM-based search). The output is a list of predicted pairs on overrepresented distances with assigned scores, P-values and plots showing the distribution of pairs in the input sequences. AVAILABILITY: The tool is available at https://www.omnifung.hki-jena.de/Rpad/Distance_Scan/index.htm.
Subject(s)
Promoter Regions, Genetic , Software , Binding Sites , Databases, Genetic , Sequence Analysis, DNA/methods , Transcription Factors/metabolismABSTRACT
The Eurasian plant Stipa capillata is the most widespread species within feather grasses. Many taxa of the genus are dominants in steppe plant communities and can be used for their classification and in studies related to climate change. Moreover, some species are of economic importance mainly as fodder plants and can be used for soil remediation processes. Although large-scale molecular data has begun to appear, there is still no complete or draft genome for any Stipa species. Thus, here we present a single-molecule long-read sequencing dataset generated using the Pacific Biosciences Sequel System. A draft genome of about 1004 Mb was obtained with a contig N50 length of 351 kb. Importantly, here we report 81,224 annotated protein-coding genes, present 77,614 perfect and 58 unique imperfect SSRs, reveal the putative allopolyploid nature of S. capillata, investigate the evolutionary history of the genus, demonstrate structural heteroplasmy of the chloroplast genome and announce for the first time the mitochondrial genome in Stipa. The assembled nuclear, mitochondrial and chloroplast genomes provide a significant source of genetic data for further works on phylogeny, hybridisation and population studies within Stipa and the grass family Poaceae.