ScbR- and ScbR2-mediated signal transduction networks coordinate complex physiological responses in Streptomyces coelicolor

Abstract In model organism Streptomyces coelicolor, γ-butyrolactones (GBLs) and antibiotics were recognized as signalling molecules playing fundamental roles in intra- and interspecies communications. To dissect the GBL and antibiotic signalling networks systematically, the in vivo targets of their respective receptors ScbR and ScbR2 were identified on a genome scale by ChIP-seq. These identified targets encompass many that are known to play important roles in diverse cellular processes (e.g. gap1, pyk2, afsK, nagE2, cdaR, cprA, cprB, absA1, actII-orf4, redZ, atrA, rpsL and sigR) and they formed regulatory cascades, sub-networks and feedforward loops to elaborately control key metabolite processes, including primary and secondary metabolism, morphological differentiation and stress response. Moreover, interplay among ScbR, ScbR2 and other regulators revealed intricate cross talks between signalling pathways triggered by GBLs, antibiotics, nutrient availability and stress. Our work provides a global view on the specific responses that could be triggered by GBL and antibiotic signals in S. coelicolor, among which the main echo was the change of production profile of endogenous antibiotics and antibiotic signals manifested a role to enhance bacterial stress tolerance as well, shedding new light on GBL and antibiotic signalling networks widespread among streptomycetes.

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The Level of AdpA Directly Affects Expression of Developmental Genes in Streptomyces coelicolor

Journal of Bacteriology ◽

10.1128/jb.05734-11 ◽

2011 ◽

Vol 193 (22) ◽

pp. 6358-6365 ◽

Cited By ~ 29

Author(s):

Marcin Wolański ◽

Rafał Donczew ◽

Agnieszka Kois-Ostrowska ◽

Paweł Masiewicz ◽

Dagmara Jakimowicz ◽

...

Keyword(s):

Early Stage ◽

Response Regulator ◽

Streptomyces Coelicolor ◽

Streptomyces Griseus ◽

Morphological Differentiation ◽

Aerial Mycelium ◽

Content Type ◽

Protein Functions

AdpA is a key regulator of morphological differentiation inStreptomyces. In contrast toStreptomyces griseus, relatively little is known about AdpA protein functions inStreptomyces coelicolor. Here, we report for the first time the translation accumulation profile of theS. coelicoloradpA(adpASc) gene; the level ofS. coelicolorAdpA (AdpASc) increased, reaching a maximum in the early stage of aerial mycelium formation (after 36 h), and remained relatively stable for the next several hours (48 to 60 h), and then the signal intensity decreased considerably. AdpAScspecifically binds theadpAScpromoter regionin vitroandin vivo, suggesting that its expression is autoregulated; surprisingly, in contrast toS. griseus, the protein presumably acts as a transcriptional activator. We also demonstrate a direct influence of AdpAScon the expression of several genes whose products play key roles in the differentiation ofS. coelicolor: STI, a protease inhibitor; RamR, an atypical response regulator that itself activates expression of the genes for a small modified peptide that is required for aerial growth; and ClpP1, an ATP-dependent protease. The diverse influence of AdpAScprotein on the expression of the analyzed genes presumably results mainly from different affinities of AdpAScprotein to individual promoters.

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Diverse control of metabolism and other cellular processes in Streptomyces coelicolor by the PhoP transcription factor: genome-wide identification of in vivo targets

Nucleic Acids Research ◽

10.1093/nar/gks766 ◽

2012 ◽

Vol 40 (19) ◽

pp. 9543-9556 ◽

Cited By ~ 64

Author(s):

Nicholas E. E. Allenby ◽

Emma Laing ◽

Giselda Bucca ◽

Andrzej M. Kierzek ◽

Colin P. Smith

Keyword(s):

Transcription Factor ◽

Streptomyces Coelicolor ◽

Cellular Processes ◽

Genome Wide

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A genome engineering resource to uncover principles of cellular organization and tissue architecture by lipid signalling

10.1101/2020.02.03.933093 ◽

2020 ◽

Author(s):

Deepti Trivedi ◽

Vinitha CM ◽

Karishma Bisht ◽

Vishnu Janardan ◽

Awadhesh Pandit ◽

...

Keyword(s):

Human Disease ◽

Genome Engineering ◽

Cultured Cells ◽

Cellular Organization ◽

Cellular Processes ◽

Lipid Signalling ◽

Disease Biology ◽

A Genome ◽

Biochemical Analyses

SummaryPhosphoinositides (PI) are key regulators of cellular organization in eukaryotes and genes that tune PI signalling are implicated in human disease mechanisms. Biochemical analyses and studies in cultured cells have identified a large number of proteins that can mediate PI signalling. However, the role of such proteins in regulating cellular processes in vivo and development in metazoans remains to be understood. Here we describe a set of CRISPR based genome engineering tools that allow the manipulation of each of these proteins with spatial and temporal control during metazoan development. We demonstrate the use of these reagents to deplete a set of 103 proteins individually in the Drosophila eye and identify several new molecules that control eye development. Our work demonstrates the power of this resource in uncovering the molecular basis of tissue homeostasis during normal development and in human disease biology.

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The relA/spoT-Homologous Gene inStreptomyces coelicolor Encodes Both Ribosome-Dependent (p)ppGppSynthesizing and -Degrading Activities

Journal of Bacteriology ◽

10.1128/jb.180.16.4123-4132.1998 ◽

1998 ◽

Vol 180 (16) ◽

pp. 4123-4132 ◽

Cited By ~ 45

Author(s):

Oscar H. Martínez-Costa ◽

Miguel A. Fernández-Moreno ◽

Francisco Malpartida

Keyword(s):

Amino Acids ◽

Deletion Mutant ◽

Streptomyces Coelicolor ◽

Stringent Response ◽

Morphological Differentiation ◽

Antibiotic Biosynthesis ◽

Homologous Gene ◽

E Coli

ABSTRACT Streptomyces coelicolor (p)ppGpp synthetase (Rel protein) belongs to the RelA and SpoT (RelA/SpoT) family, which is involved in (p)ppGpp metabolism and the stringent response. The potential functions of the rel gene have been examined.S. coelicolor Rel has been shown to be ribosome associated, and its activity in vitro is ribosome dependent. Analysis in vivo of the active recombinant protein in well-defined Escherichia coli relA and relA/spoT mutants provides evidence thatS. coelicolor Rel, like native E. coli RelA, is functionally ribosome associated, resulting in ribosome-dependent (p)ppGpp accumulation upon amino acid deprivation. Expression of anS. coelicolor C-terminally deleted Rel, comprised of only the first 489 amino acids, catalyzes a ribosome-independent (p)ppGpp formation, in the same manner as the E. colitruncated RelA protein (1 to 455 amino acids). An E. coli relA spoT double deletion mutant transformed with S. coelicolor rel gene suppresses the phenotype associated with (p)ppGpp deficiency. However, in such a strain, arel-mediated (p)ppGpp response apparently occurs after glucose depletion, but only in the absence of amino acids. Analysis of ppGpp decay in E. coli expressing the S. coelicolor rel gene suggests that it also encodes a (p)ppGpp-degrading activity. By deletion analysis, the catalytic domains of S. coelicolor Rel for (p)ppGpp synthesis and degradation have been located within its N terminus (amino acids 267 to 453 and 93 to 397, respectively). In addition,E. coli relA in an S. coelicolor reldeletion mutant restores actinorhodine production and shows a nearly normal morphological differentiation, as does the wild-typerel gene, which is in agreement with the proposed role of (p)ppGpp nucleotides in antibiotic biosynthesis.

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Genome-Wide Mutagenesis Links Multiple Metabolic Pathways with Actinorhodin Production inStreptomyces coelicolor

Applied and Environmental Microbiology ◽

10.1128/aem.03005-18 ◽

2019 ◽

Vol 85 (7) ◽

Cited By ~ 6

Author(s):

Zhong Xu ◽

Yuanyuan Li ◽

Yemin Wang ◽

Zixin Deng ◽

Meifeng Tao

Keyword(s):

Metabolic Pathways ◽

Antibiotic Production ◽

Streptomyces Coelicolor ◽

Acid Biosynthesis ◽

Content Type ◽

Cellular Processes ◽

Genome Wide ◽

A Genome ◽

Wide Perspective ◽

Actinorhodin Production

ABSTRACTStreptomycesspecies are important antibiotic-producing organisms that tightly regulate their antibiotic production. Actinorhodin is a typical antibiotic produced by the model actinomyceteStreptomyces coelicolor. To discover the regulators of actinorhodin production, we constructed a library of 50,000 independent mutants with hyperactive Tn5transposase-based transposition systems. Five hundred fifty-one genes were found to influence actinorhodin production in 988 individual mutants. Genetic complementation suggested that most of the insertions (76%) were responsible for the changes in antibiotic production. Genes involved in diverse cellular processes such as amino acid biosynthesis, carbohydrate metabolism, cell wall homeostasis, and DNA metabolism affected actinorhodin production. Genome-wide mutagenesis can identify novel genes and pathways that impact antibiotic levels, potentially aiding in engineering strains to optimize the production of antibiotics inStreptomyces.IMPORTANCEPrevious studies have shown that various genes can influence antibiotic production inStreptomycesand that intercommunication between regulators can complicate antibiotic production. Therefore, to gain a better understanding of antibiotic regulation, a genome-wide perspective on genes that influence antibiotic production was needed. We searched for genes that affected production of the antibiotic actinorhodin using a genome-wide gene disruption system. We identified 551 genes that altered actinorhodin levels, and more than half of these genes were newly identified effectors. Some of these genes may be candidates for engineeringStreptomycesstrains to improve antibiotic production levels.

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A genome engineering resource to uncover principles of cellular organization and tissue architecture by lipid signaling

eLife ◽

10.7554/elife.55793 ◽

2020 ◽

Vol 9 ◽

Author(s):

Deepti Trivedi ◽

Vinitha CM ◽

Karishma Bisht ◽

Vishnu Janardan ◽

Awadhesh Pandit ◽

...

Keyword(s):

Human Disease ◽

Genome Engineering ◽

Cultured Cells ◽

Cellular Organization ◽

Cellular Processes ◽

Disease Biology ◽

A Genome ◽

Biochemical Analyses

Phosphoinositides (PI) are key regulators of cellular organization in eukaryotes and genes that tune PI signaling are implicated in human disease mechanisms. Biochemical analyses and studies in cultured cells have identified a large number of proteins that can mediate PI signaling. However, the role of such proteins in regulating cellular processes in vivo and development in metazoans remains to be understood. Here, we describe a set of CRISPR-based genome engineering tools that allow the manipulation of each of these proteins with spatial and temporal control during metazoan development. We demonstrate the use of these reagents to deplete a set of 103 proteins individually in the Drosophila eye and identify several new molecules that control eye development. Our work demonstrates the power of this resource in uncovering the molecular basis of tissue homeostasis during normal development and in human disease biology.

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On the Way to Understanding the Interplay between the RNA Structure and Functions in Cells: A Genome-Wide Perspective

International Journal of Molecular Sciences ◽

10.3390/ijms21186770 ◽

2020 ◽

Vol 21 (18) ◽

pp. 6770

Author(s):

Angelika Andrzejewska ◽

Małgorzata Zawadzka ◽

Katarzyna Pachulska-Wieczorek

Keyword(s):

Rna Structure ◽

Gene Expression Regulation ◽

Biological Activities ◽

Current View ◽

Cellular Processes ◽

Mrna Structure ◽

Genome Wide ◽

A Genome ◽

Wide Perspective

RNAs adopt specific structures in order to perform their biological activities. The structure of RNA is an important layer of gene expression regulation, and can impact a plethora of cellular processes, starting with transcription, RNA processing, and translation, and ending with RNA turnover. The development of high-throughput technologies has enabled a deeper insight into the sophisticated interplay between the structure of the cellular transcriptome and the living cells environment. In this review, we present the current view on the RNA structure in vivo resulting from the most recent transcriptome-wide studies in different organisms, including mammalians, yeast, plants, and bacteria. We focus on the relationship between the mRNA structure and translation, mRNA stability and degradation, protein binding, and RNA posttranscriptional modifications.

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GntR-like SCO3932 Protein Provides a Link between Actinomycete Integrative and Conjugative Elements and Secondary Metabolism

International Journal of Molecular Sciences ◽

10.3390/ijms222111867 ◽

2021 ◽

Vol 22 (21) ◽

pp. 11867

Author(s):

Krzysztof J. Pawlik ◽

Mateusz Zelkowski ◽

Mateusz Biernacki ◽

Katarzyna Litwinska ◽

Pawel Jaworski ◽

...

Keyword(s):

Secondary Metabolism ◽

Streptomyces Coelicolor ◽

Morphological Differentiation ◽

Neighboring Gene ◽

Mobility Shift ◽

Integrative And Conjugative Elements ◽

Mobility Shift Assay ◽

Metabolite Synthesis

Streptomyces bacteria produce a plethora of secondary metabolites including the majority of medically important antibiotics. The onset of secondary metabolism is correlated with morphological differentiation and controlled by a complex regulatory network involving numerous regulatory proteins. Control over these pathways at the molecular level has a medical and industrial importance. Here we describe a GntR-like DNA binding transcription factor SCO3932, encoded within an actinomycete integrative and conjugative element, which is involved in the secondary metabolite biosynthesis regulation. Affinity chromatography, electrophoresis mobility shift assay, footprinting and chromatin immunoprecipitation experiments revealed, both in vitro and in vivo, SCO3932 binding capability to its own promoter region shared with the neighboring gene SCO3933, as well as promoters of polyketide metabolite genes, such as cpkD, a coelimycin biosynthetic gene, and actII-orf4—an activator of actinorhodin biosynthesis. Increased activity of SCO3932 target promoters, as a result of SCO3932 overproduction, indicates an activatory role of this protein in Streptomyces coelicolor A3(2) metabolite synthesis pathways.

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Repression of Antibiotic Production and Sporulation in Streptomyces coelicolor by Overexpression of a TetR Family Transcriptional Regulator

Applied and Environmental Microbiology ◽

10.1128/aem.00819-10 ◽

2010 ◽

Vol 76 (23) ◽

pp. 7741-7753 ◽

Cited By ~ 24

Author(s):

Delin Xu ◽

Nicolas Seghezzi ◽

Catherine Esnault ◽

Marie-Joelle Virolle

Keyword(s):

Target Genes ◽

Antibiotic Production ◽

Streptomyces Coelicolor ◽

Regulatory Gene ◽

Morphological Differentiation ◽

Promoter Sequence ◽

Inhibitory Effect ◽

Binding Motif

ABSTRACT The overexpression of a regulatory gene of the TetR family (SCO3201) originating either from Streptomyces lividans or from Streptomyces coelicolor was shown to strongly repress antibiotic production (calcium-dependent antibiotic [CDA], undecylprodigiosin [RED], and actinorhodin [ACT]) of S. coelicolor and of the ppk mutant strain of S. lividans. Curiously, the overexpression of this gene also had a strong inhibitory effect on the sporulation process of S. coelicolor but not on that of S. lividans. SCO3201 was shown to negatively regulate its own transcription, and its DNA binding motif was found to overlap its −35 promoter sequence. The interruption of this gene in S. lividans or S. coelicolor did not lead to any obvious phenotypes, indicating that when overexpressed SCO3201 likely controls the expression of target genes of other TetR regulators involved in the regulation of the metabolic and morphological differentiation process in S. coelicolor. The direct and functional interaction of SCO3201 with the promoter region of scbA, a gene under the positive control of the TetR-like regulator, ScbR, was indeed demonstrated by in vitro as well as in vivo approaches.

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Roles of Aconitase in Growth, Metabolism, and Morphological Differentiation of Streptomyces coelicolor

Journal of Bacteriology ◽

10.1128/jb.183.10.3193-3203.2001 ◽

2001 ◽

Vol 183 (10) ◽

pp. 3193-3203 ◽

Cited By ~ 33

Author(s):

P. H. Viollier ◽

K. T. Nguyen ◽

W. Minas ◽

M. Folcher ◽

G. E. Dale ◽

...

Keyword(s):

Citrate Synthase ◽

Streptomyces Coelicolor ◽

Morphological Differentiation ◽

Tca Cycle ◽

Antibiotic Biosynthesis ◽

Complete Medium ◽

Iron Regulatory Proteins ◽

Early Growth Phase

ABSTRACT The studies of aconitase presented here, along with those of citrate synthase (P. H. Viollier, W. Minas, G. E. Dale, M. Folcher, and C. J. Thompson, J. Bacteriol. 183:3184–3192, 2001), were undertaken to investigate the role of the tricarboxylic acid (TCA) cycle in Streptomyces coelicolor development. A single aconitase activity (AcoA) was detected in protein extracts of cultures during column purification. The deduced amino acid sequence of the cloned acoA gene constituted the N-terminal sequence of semipurified AcoA and was homologous to bacterial A-type aconitases and bifunctional eukaryotic aconitases (iron regulatory proteins). The fact that an acoA disruption mutant (BZ4) did not grow on minimal glucose media in the absence of glutamate confirmed that this gene encoded the primary vegetative aconitase catalyzing flux through the TCA cycle. On glucose-based complete medium, BZ4 had defects in growth, antibiotic biosynthesis, and aerial hypha formation, partially due to medium acidification and accumulation of citrate. The inhibitory effects of acids and citrate on BZ4 were partly suppressed by buffer or by introducing a citrate synthase mutation. However, the fact that growth of an acoA citA mutant remained impaired, even on a nonacidogenic carbon source, suggested alternative functions of AcoA. Immunoblots revealed that AcoA was present primarily during substrate mycelial growth on solid medium. Transcription ofacoA was limited to the early growth phase in liquid cultures from a start site mapped in vitro and in vivo.

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