scholarly journals Caries-Associated Biosynthetic Gene Clusters in Streptococcus mutans

2020 ◽  
Vol 99 (8) ◽  
pp. 969-976 ◽  
Author(s):  
S.S. Momeni ◽  
S.M. Beno ◽  
J.L. Baker ◽  
A. Edlund ◽  
T. Ghazal ◽  
...  
Keyword(s):  
Streptococcus Mutans ◽  
Genome Sequencing ◽  
Large Scale ◽  
Biological Activities ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Microbial Diseases ◽  
High Caries Risk

Early childhood caries (ECC) is a chronic disease affecting the oral health of children globally. This disease is multifactorial, but a primary factor is cariogenic microorganisms such as Streptococcus mutans. Biosynthetic gene clusters (BGCs) encode small molecules with diverse biological activities that influence the development of many microbial diseases, including caries. The purpose of this study was to identify BGCs in S. mutans from a high-caries risk study population using whole-genome sequencing and assess their association with ECC. Forty representative S. mutans isolates were selected for genome sequencing from a large-scale epidemiological study of oral microbiology and dental caries in children from a localized Alabama population. A total of 252 BGCs were identified using the antiSMASH BGC-mining tool. Three types of BGCs identified herein—butyrolactone-like, ladderane-like, and butyrolactone-ladderane-like hybrid (BL-BGC)—have not been reported in S. mutans. These 3 BGCs were cross-referenced against public transcriptomics data, and were found to be highly expressed in caries subjects. Furthermore, based on a polymerase chain reaction screening for core BL genes, 93% of children with BL-BGC had ECC. The role of BL-BGC was further investigated by examining cariogenic traits and strain fitness in a deletion mutant using in vitro biofilm models. Deletion of the BL-BGC significantly increased biofilm pH as compared to the parent strain, while other virulence and fitness properties remained unchanged. Intriguingly, BL-BGC containing strains produced more acid, a key cariogenic feature, and less biofilm than the model cariogenic strain S. mutans UA159, suggesting the importance of this BL-BGC in S. mutans–mediated cariogenesity. The structure of any BL-BGC derived metabolites, their functions, and mechanistic connection with acid production remain to be elucidated. Nevertheless, this study is the first to report the clinical significance of a BL-BGC in S. mutans. This study also highlights pangenomic diversity, which is likely to affect phenotype and virulence.

scholarly journals Global biogeographic sampling of bacterial secondary metabolism

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Zachary Charlop-Powers ◽  
Jeremy G Owen ◽  
Boojala Vijay B Reddy ◽  
Melinda A Ternei ◽  
Denise O Guimarães ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Genome Sequencing ◽  
Geographic Distance ◽  
Gene Clusters ◽  
Natural World ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Road Map ◽  
Two Factors ◽  
Natural Products Discovery

Recent bacterial (meta)genome sequencing efforts suggest the existence of an enormous untapped reservoir of natural-product-encoding biosynthetic gene clusters in the environment. Here we use the pyro-sequencing of PCR amplicons derived from both nonribosomal peptide adenylation domains and polyketide ketosynthase domains to compare biosynthetic diversity in soil microbiomes from around the globe. We see large differences in domain populations from all except the most proximal and biome-similar samples, suggesting that most microbiomes will encode largely distinct collections of bacterial secondary metabolites. Our data indicate a correlation between two factors, geographic distance and biome-type, and the biosynthetic diversity found in soil environments. By assigning reads to known gene clusters we identify hotspots of biomedically relevant biosynthetic diversity. These observations not only provide new insights into the natural world, they also provide a road map for guiding future natural products discovery efforts.

scholarly journals An Orphan MbtH-Like Protein Interacts with Multiple Nonribosomal Peptide Synthetases inMyxococcus xanthusDK1622

2018 ◽  
Vol 200 (21) ◽  
Author(s):  
Karla J. Esquilín-Lebrón ◽  
Tye O. Boynton ◽  
Lawrence J. Shimkets ◽  
Michael G. Thomas
Keyword(s):  
Myxococcus Xanthus ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Nonribosomal Peptide ◽  
Content Type ◽  
Nonribosomal Peptide Synthetases ◽  
Peptide Synthetases

ABSTRACTOne mechanism by which bacteria and fungi produce bioactive natural products is the use of nonribosomal peptide synthetases (NRPSs). Many NRPSs in bacteria require members of the MbtH-like protein (MLP) superfamily for their solubility or function. Although MLPs are known to interact with the adenylation domains of NRPSs, the role MLPs play in NRPS enzymology has yet to be elucidated. MLPs are nearly always encoded within the biosynthetic gene clusters (BGCs) that also code for the NRPSs that interact with the MLP. Here, we identify 50 orphan MLPs from diverse bacteria. An orphan MLP is one that is encoded by a gene that is not directly adjacent to genes predicted to be involved in nonribosomal peptide biosynthesis. We targeted the orphan MLP MXAN_3118 fromMyxococcus xanthusDK1622 for characterization. TheM. xanthusDK1622 genome contains 15 NRPS-encoding BGCs but only one MLP-encoding gene (MXAN_3118). We tested the hypothesis that MXAN_3118 interacts with one or more NRPS using a combination ofin vivoandin vitroassays. We determined that MXAN_3118 interacts with at least seven NRPSs from distinct BGCs. We show that one of these BGCs codes for NRPS enzymology that likely produces a valine-rich natural product that inhibits the clumping ofM. xanthusDK1622 in liquid culture. MXAN_3118 is the first MLP to be identified that naturally interacts with multiple NRPS systems in a single organism. The finding of an MLP that naturally interacts with multiple NRPS systems suggests it may be harnessed as a “universal” MLP for generating functional hybrid NRPSs.IMPORTANCEMbtH-like proteins (MLPs) are essential accessory proteins for the function of many nonribosomal peptide synthetases (NRPSs). We identified 50 MLPs from diverse bacteria that are coded by genes that are not located near any NRPS-encoding biosynthetic gene clusters (BGCs). We define these as orphan MLPs because their NRPS partner(s) is unknown. Investigations into the orphan MLP fromMyxococcus xanthusDK1622 determined that it interacts with NRPSs from at least seven distinct BGCs. Support for these MLP-NRPS interactions came from the use of a bacterial two-hybrid assay and copurification of the MLP with various NRPSs. The flexibility of this MLP to naturally interact with multiple NRPSs led us to hypothesize that this MLP may be used as a “universal” MLP during the construction of functional hybrid NRPSs.

scholarly journals In Vitro CRISPR/Cas9 System for Efficient Targeted DNA Editing

mBio ◽  
2015 ◽  
Vol 6 (6) ◽  
Author(s):  
Yunkun Liu ◽  
Weixin Tao ◽  
Shishi Wen ◽  
Zhengyuan Li ◽  
Anna Yang ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Gene Clusters ◽  
Specific Gene ◽  
Dna Fragments ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Highly Efficient ◽  
Dna Editing

ABSTRACT The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system, an RNA-guided nuclease for specific genome editing in vivo, has been adopted in a wide variety of organisms. In contrast, the in vitro application of the CRISPR/Cas9 system has rarely been reported. We present here a highly efficient in vitro CRISPR/Cas9-mediated editing (ICE) system that allows specific refactoring of biosynthetic gene clusters in Streptomyces bacteria and other large DNA fragments. Cleavage by Cas9 of circular pUC18 DNA was investigated here as a simple model, revealing that the 3′→5′ exonuclease activity of Cas9 generates errors with 5 to 14 nucleotides (nt) randomly missing at the editing joint. T4 DNA polymerase was then used to repair the Cas9-generated sticky ends, giving substantial improvement in editing accuracy. Plasmid pYH285 and cosmid 10A3, harboring a complete biosynthetic gene cluster for the antibiotics RK-682 and holomycin, respectively, were subjected to the ICE system to delete the rkD and homE genes in frame. Specific insertion of the ampicillin resistance gene (bla) into pYH285 was also successfully performed. These results reveal the ICE system to be a rapid, seamless, and highly efficient way to edit DNA fragments, and a powerful new tool for investigating and engineering biosynthetic gene clusters. IMPORTANCE Recent improvements in cloning strategies for biosynthetic gene clusters promise rapid advances in understanding and exploiting natural products in the environment. For manipulation of such biosynthetic gene clusters to generate valuable bioactive compounds, efficient and specific gene editing of these large DNA fragments is required. In this study, a highly efficient in vitro DNA editing system has been established. When combined with end repair using T4 DNA polymerase, Cas9 precisely and seamlessly catalyzes targeted editing, including in-frame deletion or insertion of the gene(s) of interest. This in vitro CRISPR editing (ICE) system promises a step forward in our ability to engineer biosynthetic pathways.

scholarly journals Chromatin-dependent regulation of secondary metabolite biosynthesis in fungi: is the picture complete?

2019 ◽  
Author(s):  
Jérôme Collemare ◽  
Michael F Seidl
Keyword(s):  
Secondary Metabolites ◽  
Secondary Metabolite ◽  
Biological Activities ◽  
Gene Clusters ◽  
Research Field ◽  
Chromatin Modifications ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Post Translational Modifications ◽  
Fungal Secondary Metabolites

ABSTRACTFungal secondary metabolites are small molecules that exhibit diverse biological activities exploited in medicine, industry and agriculture. Their biosynthesis is governed by co-expressed genes that often co-localize in gene clusters. Most of these secondary metabolite gene clusters are inactive under laboratory conditions, which is due to a tight transcriptional regulation. Modifications of chromatin, the complex of DNA and histone proteins influencing DNA accessibility, play an important role in this regulation. However, tinkering with well-characterised chemical and genetic modifications that affect chromatin alters the expression of only few biosynthetic gene clusters, and thus the regulation of the vast majority of biosynthetic pathways remains enigmatic. In the past, attempts to activate silent gene clusters in fungi mainly focused on histone acetylation and methylation, while in other eukaryotes many other post-translational modifications are involved in transcription regulation. Thus, how chromatin regulates the expression of gene clusters remains a largely unexplored research field. In this review, we argue that focusing on only few well-characterised chromatin modifications is significantly hampering our understanding of the chromatin-based regulation of biosynthetic gene clusters. Research on underexplored chromatin modifications and on the interplay between different modifications is timely to fully explore the largely untapped reservoir of fungal secondary metabolites.

scholarly journals Zn-dependent bifunctional proteases are responsible for leader peptide processing of class III lanthipeptides

2019 ◽  
Vol 116 (7) ◽  
pp. 2533-2538 ◽  
Author(s):  
Shaoming Chen ◽  
Bing Xu ◽  
Erquan Chen ◽  
Jiaqi Wang ◽  
Jingxia Lu ◽  
...  
Keyword(s):  
Gene Clusters ◽  
Leader Peptide ◽  
Biosynthetic Gene ◽  
Class Iii ◽  
Biosynthetic Gene Clusters ◽  
Peptide Processing ◽  
Leader Peptides ◽  
Biochemical Studies ◽  
Modified Peptides

Lanthipeptides are an important subfamily of ribosomally synthesized and posttranslationally modified peptides, and the removal of their N-terminal leader peptides by a designated protease(s) is a key step during maturation. Whereas proteases for class I and II lanthipeptides are well-characterized, the identity of the protease(s) responsible for class III leader processing remains unclear. Herein, we report that the class III lanthipeptide NAI-112 employs a bifunctional Zn-dependent protease, AplP, with both endo- and aminopeptidase activities to complete leader peptide removal, which is unprecedented in the biosynthesis of lanthipeptides. AplP displays a broad substrate scope in vitro by processing a number of class III leader peptides. Furthermore, our studies reveal that AplP-like proteases exist in the genomes of all class III lanthipeptide-producing strains but are usually located outside the biosynthetic gene clusters. Biochemical studies show that AplP-like proteases are universally responsible for the leader removal of the corresponding lanthipeptides. In addition, AplP-like proteases are phylogenetically correlated with aminopeptidase N from Escherichia coli, and might employ a single active site to catalyze both endo- and aminopeptidyl hydrolysis. These findings solve the long-standing question as to the mechanism of leader peptide processing during class III lanthipeptide biosynthesis, and pave the way for the production and bioengineering of this class of natural products.

scholarly journals Large-Scale Metagenome Assembly Reveals Novel Animal-Associated Microbial Genomes, Biosynthetic Gene Clusters, and Other Genetic Diversity

mSystems ◽  
2020 ◽  
Vol 5 (6) ◽  
Author(s):  
Nicholas D. Youngblut ◽  
Jacobo de la Cuesta-Zuluaga ◽  
Georg H. Reischer ◽  
Silke Dauser ◽  
Nathalie Schuster ◽  
...  
Keyword(s):  
Large Scale ◽  
Animal Species ◽  
Gene Clusters ◽  
Genomic Diversity ◽  
Data Sets ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
Microbial Genomes ◽  
Metagenome Assembly ◽  
Gut Metagenome

ABSTRACT Large-scale metagenome assemblies of human microbiomes have produced a vast catalogue of previously unseen microbial genomes; however, comparatively few microbial genomes derive from other vertebrates. Here, we generated 5,596 metagenome-assembled genomes (MAGs) from the gut metagenomes of 180 predominantly wild animal species representing 5 classes, in addition to 14 existing animal gut metagenome data sets. The MAGs comprised 1,522 species-level genome bins (SGBs), most of which were novel at the species, genus, or family level, and the majority were enriched in host versus environment metagenomes. Many traits distinguished SGBs enriched in host or environmental biomes, including the number of antimicrobial resistance genes. We identified 1,986 diverse biosynthetic gene clusters; only 23 clustered with any MIBiG database references. Gene-based assembly revealed tremendous gene diversity, much of it host or environment specific. Our MAG and gene data sets greatly expand the microbial genome repertoire and provide a broad view of microbial adaptations to the vertebrate gut. IMPORTANCE Microbiome studies on a select few mammalian species (e.g., humans, mice, and cattle) have revealed a great deal of novel genomic diversity in the gut microbiome. However, little is known of the microbial diversity in the gut of other vertebrates. We studied the gut microbiomes of a large set of mostly wild animal species consisting of mammals, birds, reptiles, amphibians, and fish. Unfortunately, we found that existing reference databases commonly used for metagenomic analyses failed to capture the microbiome diversity among vertebrates. To increase database representation, we applied advanced metagenome assembly methods to our animal gut data and to many public gut metagenome data sets that had not been used to obtain microbial genomes. Our resulting genome and gene cluster collections comprised a great deal of novel taxonomic and genomic diversity, which we extensively characterized. Our findings substantially expand what is known of microbial genomic diversity in the vertebrate gut.

scholarly journals Genome Sequencing and analyses of Two Marine Fungi from the North Sea Unraveled a Plethora of Novel Biosynthetic Gene Clusters

2018 ◽  
Vol 8 (1) ◽  
Author(s):  
Abhishek Kumar ◽  
Jens Laurids Sørensen ◽  
Frederik Teilfeldt Hansen ◽  
Mikko Arvas ◽  
Muhammad Fahad Syed ◽  
...  
Keyword(s):  
Genome Sequencing ◽  
North Sea ◽  
Marine Fungi ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters ◽  
The North ◽  
The North Sea

scholarly journals Genomic and transcriptomic survey of an endophytic fungus Calcarisporium arbuscula NRRL 3705 and potential overview of its secondary metabolites

2019 ◽  
Author(s):  
Jintao Cheng ◽  
Fei Cao ◽  
Xinai Chen ◽  
Yongquan Li ◽  
Xuming Mao
Keyword(s):  
Secondary Metabolites ◽  
Gene Cluster ◽  
Endophytic Fungi ◽  
Single Molecule ◽  
Endophytic Fungus ◽  
Biological Activities ◽  
Housekeeping Genes ◽  
Gene Clusters ◽  
Biosynthetic Gene ◽  
Biosynthetic Gene Clusters

Abstract Endophytic fungi can produce many active secondary metabolites, which are important resources of natural medicines. However, there is currently little understanding of endophytic fungi at the omics levels. Calcarisporium arbuscula , an endophytic fungus from the healthy fruit of russulaceae, can produce a variety of secondary metabolites with anti-cancer, anti-nematode and antibiotic bioactivities. Comprehensive survey of the endophytic fungi genome and transcriptome will help to understand their capacity to biosynthesize secondary metabolites and lay the foundation for the development of these precious resources. In this study，we reported the high-quality genome sequence of a strain C. arbuscula NRRL 3705 based on Single Molecule Real-Time sequencing technology. The genome of this fungus is over 45 Mb in size, relatively larger than other typical filamentous fungi, and comprises 10,001 predictable genes, encoding at least 762 secretory-proteins, 386 carbohydrate-active enzymes and 177 P450 enzymes. 398 virulence factors and 228 genes related to pathogen-host interactions were also predicted in this fungus. Moreover , 65 secondary metabolite biosynthetic gene clusters were revealed, including the gene cluster for mycotoxins aurovertins. In addition, several gene clusters were predicted to produce various mycotoxins, including aflatoxin, alternariol, destruxin, citrinin and isoflavipucine. Notably, two independent gene clusters were shown possibly involved in the biosynthesis of alternariol. Furthermore, RNA-Seq assay showed that only the expression of aurovertin gene cluster is much stronger than the housekeeping genes under laboratory conditions, consistent with that aurovertins are the predominant metabolites. The gene expression of the remaining 64 gene clusters for compound backbone biosynthesis was all lower than the housekeeping genes, which might partially explain poor production of other secondary metabolites in this fungus.Our omics data along with bioinformatics analysis indicated that C. arbuscula NRRL 3705 contains a large number of biosynthetic gene clusters and has a huge potential to produce profound secondary metabolites. This work also provides the basis for development of endophytic fungi as a new resource of natural products with promising biological activities.

scholarly journals TaxiBGC: a Taxonomy-guided Approach for the Identification of Experimentally Verified Microbial Biosynthetic Gene Clusters in Shotgun Metagenomic Data

2021 ◽  
Author(s):  
Utpal Bakshi ◽  
Vinod K Gupta ◽  
Aileen R Lee ◽  
John M Davis ◽  
Sriram Chandrasekaran ◽  
...  
Keyword(s):  
Large Scale ◽  
Human Microbiome ◽  
Gene Clusters ◽  
Human Microbiome Project ◽  
Metagenomic Data ◽  
Biosynthetic Gene ◽  
Case Control Studies ◽  
Biosynthetic Gene Clusters ◽  
Host Interactions ◽  
Microbiome Data

Biosynthetic gene clusters (BGCs) in microbial genomes encode for the production of bioactive secondary metabolites (SMs). Given the well-recognized importance of SMs in microbe-microbe and microbe-host interactions, the large-scale identification of BGCs from microbial metagenomes could offer novel functional insights into complex chemical ecology. Despite recent progress, currently available tools for predicting BGCs from shotgun metagenomes have several limitations, including the need for computationally demanding read-assembly and prediction of a narrow breadth of BGC classes. To overcome these limitations, we developed TaxiBGC (Taxonomy-guided Identification of Biosynthetic Gene Clusters), a computational pipeline for identifying experimentally verified BGCs in shotgun metagenomes by first pinpointing the microbial species likely to produce them. We show that our species-centric approach was able to identify BGCs in simulated metagenomes more accurately than by solely detecting BGC genes. By applying TaxiBGC on 5,423 metagenomes from the Human Microbiome Project and various case-control studies, we identified distinct BGC signatures of major human body sites and candidate stool-borne biomarkers for multiple diseases, including inflammatory bowel disease, colorectal cancer, and psychiatric disorders. In all, TaxiBGC demonstrates a significant advantage over existing techniques for systematically characterizing BGCs and inferring their SMs from microbiome data.

Sign in / Sign up

Export Citation Format

Share Document