Biology subjects

Pernodet, J.-L.

Publications and source records attributed to Pernodet, J.-L..

6 recordsLinked to original sources

bioRxiv2026

Effect of gene cluster relocation to the central chromosomal compartment on its expression in Streptomyces

Streptomyces bacteria are renowned for their intricate life cycle and prolific production of specialized metabolites, including antibiotics. Their linear chromosome is spatially compartmentalized: the central region contains highly conserved and expressed genes, while the terminal regions harbor less conserved, poorly expressed sequences, often rich in specialized metabolite biosynthetic gene clusters. To investigate the relationship between genome architecture and gene expression, we relocated the congocidine antibiotic biosynthetic gene cluster (CGC) from its native terminal position to the central compartment in Streptomyces ambofaciens. This relocation enhanced CGC transcription compared to its original terminal location, both in antisense orientation during exponential growth and in sense orientation after metabolic differentiation, resulting in 50% increase in congocidine production. At the 3D-level, transcription-induced domains formed at both the relocated and native CGC sites, creating sharp boundaries at a larger scale. Notably, the formation of such a boundary in the central compartment during the early stationary phase did not disrupt interarm contacts or affect neighboring gene expression. These results indicate that relocating a terminal cluster to the central chromosomal compartment provides a more favorable environment for transcription without altering chromosome compaction in the stationary phase, offering a promising strategy to enhance antibiotic production in the native host. Key points- Central relocation of a gene cluster enhanced its transcription while preserving chromosome compaction. - A transcription-induced domain formed at the new locus without altering neighboring gene expression. - This strategy increased antibiotic yield by 50% in the native host.

microbiology↗

bioRxiv2024

Root associated Streptomyces produce galbonolides to modulate plant immunity and promote rhizosphere colonisation

The rhizosphere, which serves as the primary interface between plant roots and the soil, constitutes an ecological niche for a huge diversity of microbial communities. Currently, there is little knowledge on the nature and the function of the different metabolites released by rhizospheric microbes to facilitate colonization of this highly competitive environment. Here, we demonstrate how the production of galbonolides, a group of polyene macrolides that inhibit plant and fungal Inositol Phosphorylceramide Synthase (IPCS), empowers the rhizospheric Streptomyces strain AgN23, to thrive in the rhizosphere by triggering the plants defence mechanisms. Metabolomic analysis of AgN23-inoculated Arabidopsis roots revealed a strong induction in the production of an indole alkaloid, camalexin, which is a major phytoalexin in Arabidopsis. By using a plant mutant compromised in camalexin synthesis, we show that camalexin production is necessary for the successful colonization of the rhizosphere by AgN23. Conversely, hindering galbonolides biosynthesis in AgN23 knock-out mutant resulted in loss of inhibition of IPCS, a deficiency in plant defence activation, notably the production of camalexin, and a strongly reduced development of the mutant bacteria in the rhizosphere. Together, our results identified galbonolides as important metabolites mediating rhizosphere colonisation by Streptomyces. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=159 SRC="FIGDIR/small/576418v2_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@1ec39f3org.highwire.dtl.DTLVardef@1d0a14eorg.highwire.dtl.DTLVardef@e1f61org.highwire.dtl.DTLVardef@12e487e_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO Model summarizing the mode of action of galbonolides in stimulating plant defence to support AgN23 colonization of the rhizosphere. Galbonolides secretion by Streptomyces sp. AgN23 trigger Inositol Phosphoceramide Synthase (IPCS) inhibition in Arabidopsis root cells (orange arrow). The resulting raise in Ceramide precursors of the IPCS may result in the different defence responses associated to AgN23: Hypersensitive Responses (HR), Salicylic Acid (SA) signalling, nuclear Ca2+ influx, defence gene expression and camalexin biosynthesis. This production of camalexin (blue arrow) exert a positive effect on AgN23 growth in the rhizosphere, presumably by restricting the growth of bacterial and fungal competitors sensitive to this phytoalexin. In addition, galbonolides secretion in the rhizosphere may also directly interfere with fungal competitors of AgN23. C_FIG

microbiology↗

bioRxiv2024

SYNTERUPTOR: mining genomic islands for non-classical specialised metabolite gene clusters

Microbial specialised metabolite biosynthetic gene clusters (SMBGCs) are a formidable source of natural products of pharmaceutical interest. With the multiplication of genomic data available, very efficient bioinformatic tools for automatic SMBGC detection have been developed. Nevertheless, most of these tools identify SMBGCs based on sequence similarity with enzymes typically involved in specialised metabolism and thus may miss SMBGCs coding for under characterised enzymes. Here we present SYNTERUPTOR (https://bioi2.i2bc.paris-saclay.fr/synteruptor), a program that identifies genomic islands, known to be enriched in SMBGCs, in the genomes of closely related species. With this tool, we identified a SMBGC in the genome of Streptomyces ambofaciens ATCC23877, undetected by earlier versions of antiSMASH, and experimentally demonstrated that it directs the biosynthesis of two metabolites, one of which was identified as sphydrofuran. SYNTERUPTOR is also a valuable resource for the delineation of individual SMBGCs within antiSMASH regions that may encompass multiple clusters, and for refining the boundaries of these SMBGCs.

bioinformatics↗

bioRxiv2022

Ribosomal RNA operons define a central functional compartment in the Streptomyces chromosome

Streptomyces are prolific producers of specialized metabolites with applications in medicine and agriculture. These bacteria possess a large linear chromosome genetically compartmentalized: core genes are grouped in the central part, while terminal regions are populated by poorly conserved genes and define the chromosomal arms. In exponentially growing cells, chromosome conformation capture unveiled sharp boundaries formed by ribosomal RNA (rrn) operons that segment the chromosome into multiple domains. The first and last rrn operons delimit the highly expressed central compartment and the rather transcriptionally silent terminal compartments. Here we further explore the link between the genetic distribution of rrn operons and Streptomyces genomic compartmentalization. A large panel of genomes of species representative of the genus diversity revealed that rrn operons and core genes form a central skeleton, the former being identifiable from their core gene environment. We implemented a new nomenclature for Streptomyces genomes and trace their rrn-based evolutionary history. Remarkably, rrn operons are close to pericentric inversions. Moreover, the central compartment delimited by rrn operons has a very dense, nearly invariant core gene content. Finally, this compartment harbors genes with the highest expression levels, regardless of gene persistence and distance to the origin of replication. Our results highlight that rrn operons define the structural boundaries of a central functional compartment prone to transcription in Streptomyces.

genomics↗

bioRxiv2021

Marker-free genome engineering in Amycolatopsis using the pSAM2 site-specific recombination system

Actinobacteria belonging to the genus Amycolatopsis are important for antibiotic production and other valuable biotechnological applications such as biodegradation or bioconversion. Despite their industrial importance, tools and methods for the genetic manipulation of Amycolatopsis are less developed than in other actinobacteria such as Streptomyces. Moreover, most of the existing methods do not support convenient marker-free genome engineering. Here, we report the use of the pSAM2 site-specific recombination system for the efficient deletion of marker genes or large DNA regions in Amycolatopsis. For this purpose, we constructed a shuttle vector, replicating in Escherichia coli and Amycolatopsis, expressing the Xis and Int proteins from the Streptomyces integrative and conjugative element pSAM2. These proteins are sufficient for site-specific recombination between the attachment sites attL and attR. We also constructed two plasmids, replicative in E. coli but not in Amycolatopsis, for the integration of the recombination sites attL and attR on each side of a region targeted for deletion. We exemplified the use of these tools in Amycolatopsis mediterranei DSM 40773 by obtaining with high efficiency (>95%) a marker-free deletion of one single gene in the rifamycin biosynthetic gene cluster or of the entire 90-kb cluster. IMPORTANCEThe genus Amycolatopsis is regarded as an important source of diverse specialized metabolites. Members of this genus are used in industry for the production of valuable antibiotics such as rifamycins or vancomycin. Amycolatopsis spp. also present a great interest for biotechnological applications such as biodegradation or bioconversion. Despite their importance, their genetic manipulation was somehow hampered by the lack of efficient tools. Here we report the successful use of the pSAM2 site-specific recombination system to construct unmarked deletion mutants, allowing marker recycling, or to create large deletions in A. mediterranei DSM 40773. The high efficiency of this site-specific recombination system and it possible application to other Amycolatopsis species open new opportunities for marker-free genome engineering in this genus.

molecular biology↗

bioRxiv2020

Dynamics of the compartmentalized Streptomyces chromosome during metabolic differentiation

Streptomyces are among the most prolific bacterial producers of specialized metabolites, including antibiotics. The linear chromosome is partitioned into a central region harboring core genes and two extremities enriched in specialized metabolite biosynthetic gene clusters (SMBGCs). The molecular mechanisms governing structure and function of these compartmentalized genomes remain mostly unknown. Here we show that in exponential phase, chromosome structure correlates with genetic compartmentalization: conserved, large and highly transcribed genes form boundaries that segment the central part of the chromosome into domains, whereas the terminal ends are transcriptionally, largely quiescent compartments with different structural features. Onset of metabolic differentiation is accompanied by remodeling of chromosome architecture from an open to a rather closed conformation, in which the SMBGCs are expressed forming new boundaries. Altogether, our results reveal that S. ambofaciens linear chromosome is partitioned into structurally distinct entities, indicating a link between chromosome folding, gene expression and genome evolution.

microbiology↗