Advances in microbial culturing conditions to activate silent biosynthetic gene clusters for novel metabolite production

ABSTRACT Microbes produce a plethora of secondary (or specialized) metabolites that, although not essential for primary metabolism, benefit them to survive in the environment, communicate, and influence cell differentiation. Biosynthetic gene clusters (BGCs), responsible for the production of these secondary metabolites, are readily identifiable on bacterial genome sequences. Understanding the phylogeny and distribution of BGCs helps us to predict the natural product synthesis ability of new isolates. Here, we examined 310 genomes from the Bacillus subtilis group, determined the inter- and intraspecies patterns of absence/presence for all BGCs, and assigned them to defined gene cluster families (GCFs). This allowed us to establish patterns in the distribution of both known and unknown products. Further, we analyzed variations in the BGC structures of particular families encoding natural products, such as plipastatin, fengycin, iturin, mycosubtilin, and bacillomycin. Our detailed analysis revealed multiple GCFs that are species or clade specific and a few others that are scattered within or between species, which will guide exploration of the chemodiversity within the B. subtilis group. Surprisingly, we discovered that partial deletion of BGCs and frameshift mutations in selected biosynthetic genes are conserved within phylogenetically related isolates, although isolated from around the globe. Our results highlight the importance of detailed genomic analysis of BGCs and the remarkable phylogenetically conserved erosion of secondary metabolite biosynthetic potential in the B. subtilis group. IMPORTANCE Members of the B. subtilis species complex are commonly recognized producers of secondary metabolites, among those, the production of antifungals, which makes them promising biocontrol strains. While there are studies examining the distribution of well-known secondary metabolites in Bacilli, intraspecies clade-specific distribution has not been systematically reported for the B. subtilis group. Here, we report the complete biosynthetic potential within the B. subtilis group to explore the distribution of the biosynthetic gene clusters and to reveal an exhaustive phylogenetic conservation of secondary metabolite production within Bacillus that supports the chemodiversity within this species complex. We identify that certain gene clusters acquired deletions of genes and particular frameshift mutations, rendering them inactive for secondary metabolite biosynthesis, a conserved genetic trait within phylogenetically conserved clades of certain species. The overview guides the assignment of the secondary metabolite production potential of newly isolated Bacillus strains based on genome sequence and phylogenetic relatedness.

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Comparative genomics uncovers genetic diversity and synthetic biology of secondary metabolite production of the genus Trametes

10.21203/rs.2.11205/v1 ◽

2019 ◽

Author(s):

Yan Zhang ◽

Jie Cao ◽

Jingjing Wang ◽

Yajun Cheng ◽

Minghui Zhou ◽

...

Keyword(s):

Genetic Diversity ◽

Synthetic Biology ◽

Secondary Metabolite ◽

Gene Clusters ◽

Glycoside Hydrolases ◽

Secondary Metabolite Production ◽

Crystalline Cellulose ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Metabolite Production

Abstract It’s well-established that the CAZyme genes of genus Trametes contributed to the degradation processes of polysaccharides, including lignin or crystalline cellulose. However, the comprehensive analysis of the composition of CAZymes and the biosynthetic gene clusters of Trametes genus remain unclear. We conducted comparative analysis, detected the CAZyme genes, and predicted the biosynthetic gene clusters for 9 Trametes strains. Among 82,053 homologous clusters we obtained for genus Trametes, we identified 8,518 core genes, 60,441 accessory genes and 13,094 specific genes. Our results showed that a large proportion of CAZyme genes were catalogued into glycoside hydrolases, glycosyltransferases, and carbohydrate esterases. The predicted BGCs of Trametes genus were divided into 6 strategies and the 9 Trametes strains harbored 47.78 BGCs on average. Our study uncovers the genus Trametes exhibited an open pan-genome structure, provides insights into the genetic diversity and explores the synthetic biology of secondary metabolite production for Trametes genus.

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Transcription Factor Repurposing Offers Insights into Evolution of Biosynthetic Gene Cluster Regulation

mBio ◽

10.1128/mbio.01399-21 ◽

2021 ◽

Author(s):

Wenjie Wang ◽

Milton Drott ◽

Claudio Greco ◽

Dianiris Luciano-Rosario ◽

Pinmei Wang ◽

...

Keyword(s):

Transcription Factor ◽

Secondary Metabolites ◽

Gene Cluster ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

The Other ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Metabolite Production ◽

Fungal Secondary Metabolites

Fungal secondary metabolites (SMs) are an important source of pharmaceuticals on one hand and toxins on the other. Efforts to identify the biosynthetic gene clusters (BGCs) that synthesize SMs have yielded significant insights into how variation in the genes that compose BGCs may impact subsequent metabolite production within and between species.

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Boosting Secondary Metabolite Production and Discovery through the Engineering of Novel Microbial Biosensors

BioMed Research International ◽

10.1155/2018/7021826 ◽

2018 ◽

Vol 2018 ◽

pp. 1-11 ◽

Cited By ~ 6

Author(s):

Ulysses Amancio de Frias ◽

Greicy Kelly Bonifacio Pereira ◽

María-Eugenia Guazzaroni ◽

Rafael Silva-Rocha

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolite ◽

Gene Clusters ◽

Secondary Metabolite Production ◽

Special Focus ◽

Biosynthetic Gene ◽

Expression Systems ◽

Biosynthetic Gene Clusters ◽

Metabolite Production ◽

Synthetic Promoters

Bacteria are a source of a large number of secondary metabolites with several biomedical and biotechnological applications. In recent years, there has been tremendous progress in the development of novel synthetic biology approaches both to increase the production rate of secondary metabolites of interest in native producers and to mine and reconstruct novel biosynthetic gene clusters in heterologous hosts. Here, we present the recent advances toward the engineering of novel microbial biosensors to detect the synthesis of secondary metabolites in bacteria and in the development of synthetic promoters and expression systems aiming at the construction of microbial cell factories for the production of these compounds. We place special focus on the potential of Gram-negative bacteria as a source of biosynthetic gene clusters and hosts for pathway assembly, on the construction and characterization of novel promoters for native hosts, and on the use of computer-aided design of novel pathways and expression systems for secondary metabolite production. Finally, we discuss some of the potentials and limitations of the approaches that are currently being developed and we highlight new directions that could be addressed in the field.

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Comparative Genomics Determines Strain-Dependent Secondary Metabolite Production in Streptomyces venezuelae Strains

Biomolecules ◽

10.3390/biom10060864 ◽

2020 ◽

Vol 10 (6) ◽

pp. 864

Author(s):

Woori Kim ◽

Namil Lee ◽

Soonkyu Hwang ◽

Yongjae Lee ◽

Jihun Kim ◽

...

Keyword(s):

Secondary Metabolites ◽

Secondary Metabolite ◽

Gene Clusters ◽

Secondary Metabolite Production ◽

Biosynthetic Gene ◽

Genome Sequences ◽

Biosynthetic Gene Clusters ◽

Streptomyces Venezuelae ◽

Metabolite Production ◽

Non Coding Rna

Streptomyces venezuelae is well known to produce various secondary metabolites, including chloramphenicol, jadomycin, and pikromycin. Although many strains have been classified as S. venezuelae species, only a limited number of strains have been explored extensively for their genomic contents. Moreover, genomic differences and diversity in secondary metabolite production between the strains have never been compared. Here, we report complete genome sequences of three S. venezuelae strains (ATCC 10712, ATCC 10595, and ATCC 21113) harboring chloramphenicol and jadomycin biosynthetic gene clusters (BGC). With these high-quality genome sequences, we revealed that the three strains share more than 85% of total genes and most of the secondary metabolite biosynthetic gene clusters (smBGC). Despite such conservation, the strains produced different amounts of chloramphenicol and jadomycin, indicating differential regulation of secondary metabolite production at the strain level. Interestingly, antagonistic production of chloramphenicol and jadomycin was observed in these strains. Through comparison of the chloramphenicol and jadomycin BGCs among the three strains, we found sequence variations in many genes, the non-coding RNA coding regions, and binding sites of regulators, which affect the production of the secondary metabolites. We anticipate that these genome sequences of closely related strains would serve as useful resources for understanding the complex secondary metabolism and for designing an optimal production process using Streptomyces strains.

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Chimeric LuxR Transcription Factors Rewire Natural Product Regulation

10.1101/584987 ◽

2019 ◽

Author(s):

Ruchira Mukherji ◽

Somak Chowdhury ◽

Pierre Stallforth

Keyword(s):

Natural Product ◽

Gene Clusters ◽

Product Formation ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Activator Proteins ◽

Product Regulation ◽

Powerful Approach ◽

Culturing Conditions ◽

Modular Nature

LuxR-type transcriptional activator proteins frequently flank bacterial biosynthetic gene clusters (BGCs) where they play a crucial role in regulating natural product formation. Only few bacterial BGCs are expressed under standard culturing conditions, thus modulation of flanking LuxRs is a powerful approach to activate silent clusters. Here, we show that exploiting the modular nature LuxR proteins and constructing chimeric LuxRs enables the activation of BGCs.

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Impacts of environmental stress on growth, secondary metabolite biosynthetic gene clusters and metabolite production of xerotolerant/xerophilic fungi

Current Genetics ◽

10.1007/s00294-014-0455-9 ◽

2014 ◽

Vol 61 (3) ◽

pp. 325-334 ◽

Cited By ~ 42

Author(s):

Angel Medina ◽

Markus Schmidt-Heydt ◽

Alicia Rodríguez ◽

Roberto Parra ◽

Rolf Geisen ◽

...

Keyword(s):

Environmental Stress ◽

Secondary Metabolite ◽

Gene Clusters ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Metabolite Production

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Vertical Inheritance Facilitates Interspecies Diversification in Biosynthetic Gene Clusters and Specialized Metabolites

mBio ◽

10.1128/mbio.02700-21 ◽

2021 ◽

Author(s):

Alexander B. Chase ◽

Douglas Sweeney ◽

Mitchell N. Muskat ◽

Dulce G. Guillén-Matus ◽

Paul R. Jensen

Keyword(s):

Natural Products ◽

Gene Clusters ◽

Species Level ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Evolutionary Processes ◽

Vertical Inheritance ◽

Metabolite Production ◽

Specialized Metabolites ◽

Microbial Natural Products

Microbial natural products are traditionally exploited for their pharmaceutical potential, yet our understanding of the evolutionary processes driving BGC evolution and compound diversification remains poorly developed. While HGT is recognized as an integral driver of BGC distributions, we find that the effects of vertical inheritance on BGC diversification had direct implications for species-level specialized metabolite production.

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Phylogenetic distribution of secondary metabolites in the Bacillus subtilis species complex

10.1101/2020.10.28.358507 ◽

2020 ◽

Author(s):

Kat Steinke ◽

Omkar S. Mohite ◽

Tilmann Weber ◽

Ákos T. Kovács

Keyword(s):

Bacillus Subtilis ◽

Secondary Metabolites ◽

Detailed Analysis ◽

Secondary Metabolite ◽

Genome Sequence ◽

Species Complex ◽

Gene Clusters ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Metabolite Production

ABSTRACTMicrobes produce a plethora of secondary metabolites that although not essential for primary metabolism benefit them to survive in the environment, communicate, and influence differentiation. Biosynthetic gene clusters (BGCs) responsible for the production of these secondary metabolites are readily identifiable on the genome sequence of bacteria. Understanding the phylogeny and distribution of BGCs helps us to predict natural product synthesis ability of new isolates. Here, we examined the inter- and intraspecies patterns of absence/presence for all BGCs identified with antiSMASH 5.0 in 310 genomes from the B. subtilis group and assigned them to defined gene cluster families (GCFs). This allowed us to establish patterns in distribution for both known and unknown products. Further, we analyzed variations in the BGC structure of particular families encoding for natural products such as plipastatin, fengycin, iturin, mycosubtilin and bacillomycin. Our detailed analysis revealed multiple GCFs that are species or clade specific and few others that are scattered within or between species, which will guide exploration of the chemodiversity within the B. subtilis group. Uniquely, we discovered that partial deletion of BGCs and frameshift mutations in selected biosynthetic genes are conserved within phylogenetically related isolates, although isolated from around the globe. Our results highlight the importance of detailed analysis of BGCs and the remarkable phylogenetically conserved errodation of secondary metabolite biosynthetic potential in the B. subtilis group.IMPORTANCEMembers of the B. subtilis species complex are commonly recognized producers of secondary metabolites, among those the production of antifungals makes them promising biocontrol strains. However, while there are studies examining the distribution of well-known B. subtilis metabolites, this has not yet been systematically reported for the group. Here, we report the complete biosynthetic potential within the Bacillus subtilis group species to explore the distribution of the biosynthetic gene clusters and to provide an exhaustive phylogenetic conservation of secondary metabolite production supporting the chemodiversity of Bacilli. We identify that certain gene clusters acquired deletions of genes and particular frame-shift mutations rendering them inactive for secondary metabolite biosynthesis, a conserved genetic trait within phylogenetically conserved clades of certain species. The overview presented will superbly guide assigning the secondary metabolite production potential of newly isolated strains based on genome sequence and phylogenetic relatedness.

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The histone deacetylase clr3 regulates secondary metabolite production and growth under oxidative stress conditions in Penicillium brasilianum

10.1101/2020.05.01.072108 ◽

2020 ◽

Cited By ~ 1

Author(s):

Daniel Yuri Akiyama ◽

Marina Campos Rocha ◽

Jonas Henrique Costa ◽

Iran Malavazi ◽

Taícia Pacheco Fill

Keyword(s):

Oxidative Stress ◽

Histone Deacetylase ◽

Secondary Metabolite ◽

Filamentous Fungi ◽

Gene Clusters ◽

Stress Conditions ◽

Biosynthetic Gene ◽

Biosynthetic Gene Clusters ◽

Metabolite Production ◽

Penicillium Brasilianum

ABSTRACTMost of the biosynthetic gene clusters (BGCs) found in filamentous fungi are silent under standard laboratory cultivation conditions due to the lack of expression triggering stimuli, representing a considerable drawback in drug discovery. To access the full biosynthetic potential of these microbes, studies towards the activation of cryptic BGCs are essential. Histone acetylation status is an important regulator of chromatin structure which impacts in cell physiology and, therefore, expression of biosynthetic gene clusters in filamentous fungi. Histone deacetylases (HDACs) and histone acetyl-transferases (HATs) are responsible for maintaining and controlling this process under different cell conditions. In this study, clr3, a gene encoding a histone deacetylase in Penicillium brasilianum was deleted and associated phenotypic and metabolic changes evaluated. Results indicate reduced growth under oxidative stress conditions in the Δclr3 knockout strain. Also, the production of several secondary metabolites including austin-related meroterpenoids, brasiliamides, mycotoxins such as verruculogen and penicillic acid, as well as cyclodepsipeptides was reduced in the Δclr3 strain when compared to wild-type strain. Accordingly, addition of epigenetic modulators responsible for HDAC inhibition such as suberoylanilide hydroxamic acid (SAHA) and nicotinamide (NAA) to P. brasilianum growth media also culminated in reduction of secondary metabolite production. Mass Spectrometry Imaging (MSI) was applied to compare metabolite production and spatial distribution on the colony. Results suggest that Clr3 plays an important role in secondary metabolite biosynthesis in P. brasilianum, thus offering new strategies for regulation of natural product synthesis by assessing chromatin modification in P. brasilianum.

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