The discovery of interleukin (IL)-6 and its receptor subunits provided a foundation to understand the biology of a group of related cytokines: IL-12, IL-23, and IL-27. These family members utilize ...shared receptors and cytokine subunits and influence the outcome of cancer, infection, and inflammatory diseases. Consequently, many facets of their biology are being therapeutically targeted. Here, we review the landmark discoveries in this field, the combinatorial biology inherent to this family, and how patient datasets have underscored the critical role of these pathways in human disease. We present significant knowledge gaps, including how similar signals from these cytokines can mediate distinct outcomes, and discuss how a better understanding of the biology of the IL-12 family provides new therapeutic opportunities.
The IL-12 family of cytokines influences the outcome of cancer, infection, and inflammatory diseases. Stumhofer, Hunter, and Tait review the combinatorial biology inherent to this family and the recent clinical insights that underscore the critical role of these pathways in human disease.
Natural product-encoding biosynthetic gene clusters (BGCs) within microbial genomes far outnumber the known natural products; chemical products from such BGCs remain cryptic. These silent BGCs hold ...promise not only for the elaboration of new natural products but also for the discovery of useful biosynthetic enzymes. Here, we describe a genome mining strategy targeted toward the discovery of substrate promiscuous natural product biosynthetic enzymes. In the genome of the methanotrophic bacterium Methylovulum psychrotolerans Sph1T, we discover a transcriptionally silent natural product BGC that encoded numerous ribosomally synthesized and post-translationally modified peptide (RiPP) natural products. These cryptic RiPP natural products were accessed using heterologous expression of the substrate peptide and biosynthetic enzyme-encoded genes. In line with our genome mining strategy, the RiPP biosynthetic enzymes in this BGC were found to be substrate promiscuous, which allowed us to use them in a combinatorial fashion with a similarly substrate-tolerant cyanobactin biosynthetic enzyme to introduce head-to-tail macrocyclization in the proteusin family of RiPP natural products.
Nonribosomal peptides and polyketides are a diverse group of natural products with complex chemical structures and enormous pharmaceutical potential. They are synthesized on modular nonribosomal ...peptide synthetase (NRPS) and polyketide synthase (PKS) enzyme complexes by a conserved thiotemplate mechanism. Here, we report the widespread occurrence of NRPS and PKS genetic machinery across the three domains of life with the discovery of 3,339 gene clusters from 991 organisms, by examining a total of 2,699 genomes. These gene clusters display extraordinarily diverse organizations, and a total of 1,147 hybrid NRPS/PKS clusters were found. Surprisingly, 10% of bacterial gene clusters lacked modular organization, and instead catalytic domains were mostly encoded as separate proteins. The finding of common occurrence of nonmodular NRPS differs substantially from the current classification. Sequence analysis indicates that the evolution of NRPS machineries was driven by a combination of common descent and horizontal gene transfer. We identified related siderophore NRPS gene clusters that encoded modular and nonmodular NRPS enzymes organized in a gradient. A higher frequency of the NRPS and PKS gene clusters was detected from bacteria compared with archaea or eukarya. They commonly occurred in the phyla of Proteobacteria, Actinobacteria, Firmicutes, and Cyanobacteria in bacteria and the phylum of Ascomycota in fungi. The majority of these NRPS and PKS gene clusters have unknown end products highlighting the power of genome mining in identifying novel genetic machinery for the biosynthesis of secondary metabolites.
Abstract
Microorganisms produce small bioactive compounds as part of their secondary or specialised metabolism. Often, such metabolites have antimicrobial, anticancer, antifungal, antiviral or other ...bio-activities and thus play an important role for applications in medicine and agriculture. In the past decade, genome mining has become a widely-used method to explore, access, and analyse the available biodiversity of these compounds. Since 2011, the ‘antibiotics and secondary metabolite analysis shell—antiSMASH’ (https://antismash.secondarymetabolites.org/) has supported researchers in their microbial genome mining tasks, both as a free to use web server and as a standalone tool under an OSI-approved open source licence. It is currently the most widely used tool for detecting and characterising biosynthetic gene clusters (BGCs) in archaea, bacteria, and fungi. Here, we present the updated version 7 of antiSMASH. antiSMASH 7 increases the number of supported cluster types from 71 to 81, as well as containing improvements in the areas of chemical structure prediction, enzymatic assembly-line visualisation and gene cluster regulation.
Graphical Abstract
Graphical Abstract
antiSMASH 7 increases the number of supported cluster types from 71 to 81, as well as containing improvements in the areas of chemical structure prediction, enzymatic assembly-line visualisation and gene cluster regulation.
With an ever-increasing amount of (meta)genomic data being deposited in sequence databases, (meta)genome mining for natural product biosynthetic pathways occupies a critical role in the discovery of ...novel pharmaceutical drugs, crop protection agents and biomaterials. The genes that encode these pathways are often organised into biosynthetic gene clusters (BGCs). In 2015, we defined the Minimum Information about a Biosynthetic Gene cluster (MIBiG): a standardised data format that describes the minimally required information to uniquely characterise a BGC. We simultaneously constructed an accompanying online database of BGCs, which has since been widely used by the community as a reference dataset for BGCs and was expanded to 2021 entries in 2019 (MIBiG 2.0). Here, we describe MIBiG 3.0, a database update comprising large-scale validation and re-annotation of existing entries and 661 new entries. Particular attention was paid to the annotation of compound structures and biological activities, as well as protein domain selectivities. Together, these new features keep the database up-to-date, and will provide new opportunities for the scientific community to use its freely available data, e.g. for the training of new machine learning models to predict sequence-structure-function relationships for diverse natural products. MIBiG 3.0 is accessible online at https://mibig.secondarymetabolites.org/.
Choline and trimethylamine (TMA) are small molecules that play central roles in biological processes throughout all kingdoms of life. These ubiquitous metabolites are linked through a single ...biochemical transformation, the conversion of choline to TMA by anaerobic microorganisms. This metabolic activity, which contributes to methanogenesis and human disease, has been known for over a century but has eluded genetic and biochemical characterization. We have identified a gene cluster responsible for anaerobic choline degradation within the genome of a sulfate-reducing bacterium and verified its function using both a genetic knockout strategy and heterologous expression in Escherichia coli . Bioinformatics and electron paramagnetic resonance (EPR) spectroscopy revealed the involvement of a C–N bond cleaving glycyl radical enzyme in TMA production, which is unprecedented chemistry for this enzyme family. Our discovery provides the predictive capabilities needed to identify choline utilization clusters in numerous bacterial genomes, underscoring the importance and prevalence of this metabolic activity within the human microbiota and the environment.
Abstract
Fueled by the explosion of (meta)genomic data, genome mining of specialized metabolites has become a major technology for drug discovery and studying microbiome ecology. In these efforts, ...computational tools like antiSMASH have played a central role through the analysis of Biosynthetic Gene Clusters (BGCs). Thousands of candidate BGCs from microbial genomes have been identified and stored in public databases. Interpreting the function and novelty of these predicted BGCs requires comparison with a well-documented set of BGCs of known function. The MIBiG (Minimum Information about a Biosynthetic Gene Cluster) Data Standard and Repository was established in 2015 to enable curation and storage of known BGCs. Here, we present MIBiG 2.0, which encompasses major updates to the schema, the data, and the online repository itself. Over the past five years, 851 new BGCs have been added. Additionally, we performed extensive manual data curation of all entries to improve the annotation quality of our repository. We also redesigned the data schema to ensure the compliance of future annotations. Finally, we improved the user experience by adding new features such as query searches and a statistics page, and enabled direct link-outs to chemical structure databases. The repository is accessible online at https://mibig.secondarymetabolites.org/.
A gene cluster encoding the biosynthesis of the fungal tropolone stipitatic acid was discovered in Talaromyces stipitatus (Penicillium stipitatum) and investigated by targeted gene knockout. A ...minimum of three genes are required to form the tropolone nucleus: tropA encodes a nonreducing polyketide synthase which releases 3-methylorcinaldehyde; tropB encodes a FAD-dependent monooxygenase which dearomatizes 3-methylorcinaldehyde via hydroxylation at C-3; and tropC encodes a non-heme Fe(II)-dependent dioxygenase which catalyzes the oxidative ring expansion to the tropolone nucleus via hydroxylation of the 3-methyl group. The tropA gene was characterized by heterologous expression in Aspergillus oryzae, whereas tropB and tropC were successfully expressed in Escherichia coli and the purified TropB and TropC proteins converted 3-methylorcinaldehyde to a tropolone in vitro. Finally, knockout of the tropD gene, encoding a cytochrome P450 monooxygenase, indicated its place as the next gene in the pathway, probably responsible for hydroxylation of the 6-methyl group. Comparison of the T. stipitatus tropolone biosynthetic cluster with other known gene clusters allows clarification of important steps during the biosynthesis of other fungal compounds including the xenovulenes, citrinin, sepedonin, sclerotiorin, and asperfuranone.
The glaucous appearance of wheat (Triticum aestivum) and barley (Hordeum vulgare) plants, that is the light bluish-gray look of flag leaf, stem, and spike surfaces, results from deposition of ...cuticular β-diketone wax on their surfaces; this phenotype is associated with high yield, especially under drought conditions. Despite extensive genetic and biochemical characterization, the molecular genetic basis underlying the biosynthesis of β-diketones remains unclear. Here, we discovered that the wheat W1 locus contains a metabolic gene cluster mediating β-diketone biosynthesis. The cluster comprises genes encoding proteins of several families including type-III polyketide synthases, hydrolases, and cytochrome P450s related to known fatty acid hydroxylases. The cluster region was identified in both genetic and physical maps of glaucous and glossy tetraploid wheat, demonstrating entirely different haplotypes in these accessions. Complementary evidence obtained through gene silencing in planta and heterologous expression in bacteria supports a model for a β-diketone biosynthesis pathway involving members of these three protein families. Mutations in homologous genes were identified in the barley eceriferum mutants defective in β-diketone biosynthesis, demonstrating a gene cluster also in the β-diketone biosynthesis Cer-cqu locus in barley. Hence, our findings open new opportunities to breed major cereal crops for surface features that impact yield and stress response.
The biosynthetic gene cluster of atlantinone B (10) was discovered in Penicillium chrysogenum MT-40. A multifunctional cytochrome P450 (AtlD) encoded by the cluster is responsible for the formation ...of the unique lactone-bridged ring and the 16β-hydroxyl of atlantinone B, and a new terpene cyclase (AtlC) can unprecedentedly accept the demethylated substrate epoxyfarnesyl-DMOA (4a) to generate three bicyclic meroterpenoids (5a–5c). This study paves the way for combinatorial synthesis of structurally diverse meroterpenoids for drug discovery.
PDF
