Project description:The present work aimed at providing noval reference genes (RGs) for streptomycetes by global quantitative analysis of gene expression profile. By using the time-series microarray data obtained in different culture medium (SMM in the present work and modified R5 from publication), the stably expressed genes of S. coelicolor were screened. Further statistical, bioinformatic and biological function analysis picked out 13 candidate RGs. According to qRT-PCR assays, 5 genes with high stability were selected and used for validation in other streptomycetes to assess their prevalence. Additionally, the absolute gene expression level reflcted by RNA-seq was also taken into consideration in the present work to guide the appropriate RGs selection for streptomycetes.
Project description:The present work aimed at providing noval reference genes (RGs) for streptomycetes by global quantitative analysis of gene expression profile. By using the time-series microarray data obtained in different culture medium (SMM in the present work and modified R5 from publication), the stably expressed genes of S. coelicolor were screened. Further statistical, bioinformatic and biological function analysis picked out 13 candidate RGs. According to qRT-PCR assays, 5 genes with high stability were selected and used for validation in other streptomycetes to assess their prevalence. Additionally, the absolute gene expression level reflcted by RNA-seq was also taken into consideration in the present work to guide the appropriate RGs selection for streptomycetes.
Project description:The present work aimed at providing noval reference genes (RGs) for streptomycetes by global quantitative analysis of gene expression profile. By using the time-series microarray data obtained in different culture medium (SMM in the present work and modified R5 from publication), the stably expressed genes of S. coelicolor were screened. Further statistical, bioinformatic and biological function analysis picked out 13 candidate RGs. According to qRT-PCR assays, 5 genes with high stability were selected and used for validation in other streptomycetes to assess their prevalence. Additionally, the absolute gene expression level reflcted by RNA-seq was also taken into consideration in the present work to guide the appropriate RGs selection for streptomycetes. Transcriptional profiling of S. coelicolor M145 at different growth phases (exponential, transitional and stationary phase)
Project description:The present work aimed at providing noval reference genes (RGs) for streptomycetes by global quantitative analysis of gene expression profile. By using the time-series microarray data obtained in different culture medium (SMM in the present work and modified R5 from publication), the stably expressed genes of S. coelicolor were screened. Further statistical, bioinformatic and biological function analysis picked out 13 candidate RGs. According to qRT-PCR assays, 5 genes with high stability were selected and used for validation in other streptomycetes to assess their prevalence. Additionally, the absolute gene expression level reflcted by RNA-seq was also taken into consideration in the present work to guide the appropriate RGs selection for streptomycetes. Transcriptional profiling variation of S. coelicolor M145, M-NM-^TscbR2 mutation with jadomycin addition at 30 h.
Project description:The Streptomyces lividans lsp gene encodes a type II signal peptidase (Lsp) that cleaves the type II leader peptides of lipoproteins. Transcriptional profiling of the bacterium depleted of the lsp gene mainly resulted in deactivation of the sigma U regulon, as well as in downregulation of genes involved in the biogenesis and function of ribosomes and genes encoding some major secretory proteins as determined by hybridisation of commercially available S. lividans genome-wide microarrays. Almost 50% of the dowregulated genes have been described as forming part of the stringent response in streptomycetes. The gene encoding the S. lividans extracellular foldase, the lipoprotein FkpA, is equally downregulated. Therefore, the deletion of lsp from the S. livdans genome temporarily triggers a cellular stress where the stringent response is, at least, partially induced.
Project description:Streptomycetes are saprophytic bacteria that grow on complex polysaccharides, such as cellulose, starch, chitin and chitosan. For the monomeric building blocks glucose, maltose and N-acetylglucosamine the metabolic pathway is well documented, but that of glucosamine (GlcN) is currently unknown. Importantly, GlcN is lethal to Streptomyces coelicolor nagB mutants, which lack glucosamine-6-phosphate deaminase activity. Here we report that spontaneous and directed mutations in the gene for the ROK-family protein RokL6 (SCO1447) relieve GlcN toxicity in nagB mutants of S. coelicolor. RNA sequencing, ChIP-Seq and over-expression studies revealed that RokL6 acts by directly repressing SCO1448, which encodes a sugar exporter; RokL6 thereby only binds to the rokL6-SCO1448 intergenic region in vivo, with consensus binding site C(T)TATCAGG - 7 nt - CCTGATAG(A). The exact transcriptional start sites for rokL6 and SCO1448 were determined using 5’RACE. RokL6 represses the transcription of both rokL6 and SCO1448 by binding to overlapping promoter sequences. Taken together, our data show that RokL6 and SCO1448 are novel GlcN-related genes, whereby RokL6 directly controls the transcription of SCO1448. The latter is a key protein in the defense of S. coelicolor against the toxicity of GlcN in a nagB-mutant background, most likely via the export of GlcN-derived toxic intermediates.
Project description:Background: During the lifetime of a fermenter culture, the soil bacterium S. coelicolor undergoes a major metabolic switch from exponential growth to antibiotic production. We have studied gene expression patterns during this switch, using a specifically designed Affymetrix genechip and a high-resolution time-series of fermenter-grown samples. Results: Surprisingly, we find that the metabolic switch actually consists of multiple finely orchestrated switching events. Strongly coherent clusters of genes show drastic changes in gene expression already many hours before the classically defined transition phase where the switch from primary to secondary metabolism was expected. The main switch in gene expression takes only 2 hours, and changes in antibiotic biosynthesis genes are delayed relative to the metabolic rearrangements. Furthermore, global variation in morphogenesis genes indicates an involvement of cell differentiation pathways in the decision phase leading up to the commitment to antibiotic biosynthesis. Conclusions: Our study provides the first detailed insights into the complex sequence of early regulatory events during and preceding the major metabolic switch in S. coelicolor, which will form the starting point for future attempts at engineering antibiotic production in a biotechnological setting. Keywords: time course
Project description:Streptomycetes are major producers of a wide variety of secondary metabolites that serve as bioactive compounds. Many secondary metabolites are produced in response to environmental signals such as biotic and abiotic stresses. In this study, we identified salinity stress as one of the stimuli activating secondary metabolism in the model Streptomyces species, Streptomyces coelicolor. Comparative metabolomics revealed overproduction of several known secondary metabolites, most notably undecylprodigiosin and coelimycin P1, in addition to their biosynthetic intermediates and derivatives as well as many unknown metabolites. Transcriptomic analysis revealed activation of diverse biological processes including potassium uptake, compatible solute production and phosphate limitation stress response through conserved and species-specific mechanisms, presumably to overcome the salinity stress. This stress response leads to activation of a variety of regulatory and metabolic pathways required for production of secondary metabolites including activation of conserved metabolic pathways for energy and substrate supply and species-specific secondary metabolites biosynthetic gene clusters. Furthermore, several promoter sequences contributing to upregulation of secondary metabolism induced by salinity stress were identified. Overall, our data show how S. coelicolor copes with the salinity stress and tailors the cellular metabolism toward secondary metabolism in a conserved and species-specific manner.
The work (proposal:https://doi.org/10.46936/10.25585/60008131) conducted by the U.S. Department of Energy Joint Genome Institute (https://ror.org/04xm1d337), a DOE Office of Science User Facility, is supported by the Office of Science of the U.S. Department of Energy operated under Contract No. DE-AC02-05CH11231.