Biology subjects

Elliot, M.

Publications and source records attributed to Elliot, M..

4 recordsLinked to original sources

bioRxiv2025

LsrL modulates Lsr2-induced chromatin structure to tune biosynthetic gene cluster regulation in Streptomyces venezuelae

Specialized biosynthetic gene clusters in Streptomyces are subject to complex regulation involving both transcriptional control and chromosome organization. The nucleoid-associated protein Lsr2 silences many of these clusters, yet how it shapes the global chromatin structure and how its conserved paralog LsrL contributes to this process remain poorly understood. In this study, we applied a multi-omics approach, combining transcriptional activity, genome-wide protein-DNA binding profiles, and three-dimensional chromosome conformation to characterize the coordination of Lsr2 and LsrL in exerting transcriptional control and genome architecture in Streptomyces venezuelae. In line with established Lsr2 functions, we find that Lsr2 sets broad transcriptional boundaries, while LsrL acts in a more context-specific manner that depends on the presence of Lsr2 and may function to reinforce or modulate Lsr2-mediated silencing. Loss of Lsr2 reshaped the chromatin landscape genome-wide, relieving its restriction on short-range contacts, triggering strong transcriptional changes and new domain boundaries near de-repressed biosynthetic gene clusters. These findings establish Lsr2 as a dominant but contextually modulated regulator whose interplay with LsrL coordinates specialized metabolism with higher-order chromosome organization.

microbiology↗

bioRxiv2024

CRISPR/dCas-mediated counter-silencing: Reprogramming dCas proteins into antagonists of xenogeneic silencers

Lsr2-like nucleoid-associated proteins function as xenogeneic silencers (XSs) inhibiting expression of horizontally acquired, AT-rich DNA in actinobacteria. Interference by transcription factors can lead to counter-silencing of XS target promoters, but relief of this repression typically requires promoter engineering. In this study, we developed a novel CRISPR/dCas-mediated counter-silencing (CRISPRcosi) approach by using nuclease-deficient dCas enzymes to counteract the Lsr2-like XS protein CgpS in Corynebacterium glutamicum or Lsr2 in Streptomyces venezuelae. Systematic in vivo reporter studies with dCas9 and dCas12a and various guide RNAs revealed effective counter-silencing of different CgpS target promoters in response to guide RNA/dCas DNA binding - independent of promoter sequence modifications. The most prominent CRISPRcosi effect was observed when targeting the CgpS nucleation site, an effect that was also seen in S. venezuelae when targeting a known Lsr2 nucleation site within the chloramphenicol biosynthesis gene cluster. Analyzing the system in C. glutamicum strains lacking the XS protein CgpS revealed varying strengths of counteracting CRISPR interference (CRISPRi) effects based on the target position and strand. Genome-wide transcriptome profiling in sgRNA/dCas9 co-expressing C. glutamicum wild-type strains revealed high counter-silencing specificity with minimal off-target effects. Thus, CRISPRcosi provides a promising strategy for the precise upregulation of XS target genes with significant potential for studying gene networks as well as for developing applications in biotechnology and synthetic biology. IMPORTANCELsr2-like nucleoid-associated proteins act as xenogeneic silencers (XSs), repressing the expression of horizontally acquired, AT-rich DNA in actinobacteria. The targets of Lsr2-like proteins are very diverse, including prophage elements, virulence gene clusters and biosynthetic gene clusters. Consequently, the targeted activation of XS target genes is of interest for fundamental research and biotechnological applications. Traditional methods for counter-silencing typically require promoter modifications. In this study, we developed a novel CRISPR/dCas-mediated counter-silencing (CRISPRcosi) approach, utilizing nuclease-deficient dCas enzymes to counteract repression by Lsr2-like proteins in Corynebacterium glutamicum and Streptomyces venezuelae. The strongest effect was observed when targeting the Lsr2 nucleation site. Genome-wide transcriptome profiling revealed high specificity with minimal off-target effects. Overall, CRISPRcosi emerges as a powerful tool for the precise activation of genes silenced by xenogeneic silencers, offering new opportunities for exploring gene networks and advancing biotechnological applications.

microbiology↗

bioRxiv2019

Transcriptional response of Streptomyces coelicolor to rapid chromosome relaxation or long-term supercoiling imbalance

Negative DNA supercoiling allows chromosome condensation within a cell and facilitates DNA unwinding, which is required for the occurrence of DNA transaction processes, i.e., DNA replication, transcription and recombination. In bacteria, changes in chromosome supercoiling impact global gene expression; however, the limited studies on the global transcriptional response have focused mostly on pathogenic species and have reported various fractions of affected genes. Furthermore, the transcriptional response to long-term supercoiling imbalance is still poorly understood. Here, we address the transcriptional response to both novobiocin-induced rapid chromosome relaxation or long-term topological imbalance, both increased and decreased supercoiling, in environmental antibiotic-producing bacteria belonging to the Streptomyces genus. During the Streptomyces complex developmental cycle, multiple copies of GC-rich linear chromosomes present in hyphal cells undergo profound topological changes, from being loosely condensed in vegetative hyphae, to being highly compacted in spores. In Streptomyces, chromosomal supercoiling changes may also be triggered by environmental stressors and have been suggested to be associated with the control of antibiotic production. Remarkably, in S. coelicolor, one of model Streptomyces species, topoisomerase I (TopA) is solely responsible for the removal of negative DNA supercoils. Using a S. coelicolor strain in which topA transcription is under the control of an inducible promoter, we generated a long-term supercoiling imbalance, enabling us to identify genes involved in the supercoiling-sensitive transcriptional response. We observed that affected genes are preferentially organized in clusters, and among them, we identified a supercoiling-hypersensitive cluster (SHC) located in the core of the S. coelicolor chromosome. Moreover, using a gyrase inhibitor, we identified the directly affected novobiocin-sensitive genes and established that the AT content in their promoter regions was increased. Notably, genes whose expression was immediately impacted by gyrase inhibition encoded products associated with membrane transport or secondary metabolite synthesis. In contrast to the novobiocin-sensitive genes, the transcripts affected by long-term topological imbalance encompassed genes encoding nucleoid-associated proteins, DNA repair proteins and transcriptional regulators, including multiple developmental regulators. Collectively, our results show that long-term supercoiling imbalance globally regulates gene transcription and has the potential to impact development, secondary metabolism and DNA repair, amongst others.

microbiology↗

bioRxiv2018

Streptomyces volatile compounds influence exploration and microbial community dynamics by altering iron availability

Bacteria and fungi produce a wide array of volatile organic compounds (VOCs), and these can act as infochemicals or as competitive tools. Recent work has shown that the VOC trimethylamine (TMA) can promote a new form of Streptomyces growth, termed exploration. Here, we report that TMA also serves to alter nutrient availability in the area surrounding exploring cultures: TMA dramatically increases the environmental pH, and in doing so, reduces iron availability. This, in turn, compromised the growth of other soil bacteria and fungi. In contrast, Streptomyces thrives in these iron-depleted niches by secreting a suite of differentially modified siderophores, and by upregulating genes associated with siderophore uptake. Further reducing iron levels by siderophore piracy, limiting siderophore uptake, or growing cultures in the presence of iron chelators, unexpectedly enhanced exploration. Our work reveals a new role for VOCs in modulating iron levels in the environment, and implies a critical role for VOCs in modulating the behaviour of microbes and the makeup of their communities.

microbiology↗