SCO6992, A Protein with β-Glucuronidase Activity, Complements a Mutation at the absR Locus and Promotes Antibiotic Biosynthesis in Streptomyces coelicolor

ABSTRACT On solid media, the reproductive growth of Streptomyces involves antibiotic biosynthesis coincident with the erection of filamentous aerial hyphae. Following cessation of growth of an aerial hypha, multiple septation occurs at the tip to form a chain of unigenomic spores. A gene, crgA, that coordinates several aspects of this reproductive growth is described. The gene product is representative of a well-conserved family of small actinomycete proteins with two C-terminal hydrophobic-potential membrane-spanning segments. In Streptomyces avermitilis, crgA is required for sporulation, and inactivation of the gene abolished most sporulation septation in aerial hyphae. Disruption of the orthologous gene in Streptomyces coelicolor indicates that whereas CrgA is not essential for sporulation in this species, during growth on glucose-containing media, it influences the timing of the onset of reproductive growth, with precocious erection of aerial hyphae and antibiotic production by the mutant. Moreover, CrgA subsequently acts to inhibit sporulation septation prior to growth arrest of aerial hyphae. Overexpression of CrgA in S. coelicolor, uncoupling any nutritional and growth phase-dependent regulation, results in growth of nonseptated aerial hyphae on all media tested, consistent with a role for the protein in inhibiting sporulation septation.

Download Full-text

A Novel Two-Component System, GluR-GluK, Involved in Glutamate Sensing and Uptake in Streptomyces coelicolor

Journal of Bacteriology ◽

10.1128/jb.00097-17 ◽

2017 ◽

Vol 199 (18) ◽

Cited By ~ 6

Author(s):

Lei Li ◽

Weihong Jiang ◽

Yinhua Lu

Keyword(s):

Signal Transduction ◽

Glutamate Uptake ◽

Streptomyces Coelicolor ◽

Signal Transduction Pathway ◽

Antibiotic Biosynthesis ◽

Transduction Pathway ◽

Two Component System ◽

Component System ◽

Content Type ◽

Two Component

ABSTRACT Two-component systems (TCSs), the predominant signal transduction pathways employed by bacteria, play important roles in physiological metabolism in Streptomyces. Here, a novel TCS, GluR-GluK (encoded by SCO5778-SCO5779), which is located divergently from the gluABCD operon encoding a glutamate uptake system, was identified as being involved in glutamate sensing and uptake as well as antibiotic biosynthesis in Streptomyces coelicolor. Under the condition of minimal medium (MM) supplemented with different concentrations of glutamate, deletion of the gluR-gluK operon (gluR-K) resulted in enhanced actinorhodin (ACT) but reduced undecylprodigiosin (RED) and yellow type I polyketide (yCPK) production, suggesting that GluR-GluK plays a differential role in antibiotic biosynthesis. Furthermore, we found that the response regulator GluR directly promotes the expression of gluABCD under the culture condition of MM with a high concentration of glutamate (75 mM). Using the biolayer interferometry assay, we demonstrated that glutamate acts as the direct signal of the histidine kinase GluK. It was therefore suggested that upon sensing high concentrations of glutamate, GluR-GluK would be activated and thereby facilitate glutamate uptake by increasing gluABCD expression. Finally, we demonstrated that the role of GluR-GluK in antibiotic biosynthesis is independent of its function in glutamate uptake. Considering the wide distribution of the glutamate-sensing (GluR-GluK) and uptake (GluABCD) module in actinobacteria, it could be concluded that the GluR-GluK signal transduction pathway involved in secondary metabolism and glutamate uptake should be highly conserved in this bacterial phylum. IMPORTANCE In this study, a novel two-component system (TCS), GluR-GluK, was identified to be involved in glutamate sensing and uptake as well as antibiotic biosynthesis in Streptomyces coelicolor. A possible GluR-GluK working model was proposed. Upon sensing high glutamate concentrations (such as 75 mM), activated GluR-GluK could regulate both glutamate uptake and antibiotic biosynthesis. However, under a culture condition of MM supplemented with low concentrations of glutamate (such as 10 mM), although GluR-GluK is activated, its activity is sufficient only for the regulation of antibiotic biosynthesis. To the best of our knowledge, this is the first report describing a TCS signal transduction pathway for glutamate sensing and uptake in actinobacteria.

Download Full-text

The RNA Polymerase Omega Factor RpoZ Is Regulated by PhoP and Has an Important Role in Antibiotic Biosynthesis and Morphological Differentiation in Streptomyces coelicolor

Applied and Environmental Microbiology ◽

10.1128/aem.00465-11 ◽

2011 ◽

Vol 77 (21) ◽

pp. 7586-7594 ◽

Cited By ~ 27

Author(s):

Fernando Santos-Beneit ◽

Mónica Barriuso-Iglesias ◽

Lorena T. Fernández-Martínez ◽

Miriam Martínez-Castro ◽

Alberto Sola-Landa ◽

...

Keyword(s):

Rna Polymerase ◽

Response Regulator ◽

Antibiotic Production ◽

Streptomyces Coelicolor ◽

Morphological Differentiation ◽

Luciferase Reporter ◽

Antibiotic Biosynthesis ◽

Binding Assays ◽

Content Type ◽

Phosphate Depletion

ABSTRACTThe RNA polymerase (RNAP) omega factor (ω) forms a complex with the α2ββ′ core of this enzyme in bacteria. We have characterized therpoZgene ofStreptomyces coelicolor, which encodes a small protein (90 amino acids) identified as the omega factor. Deletion of therpoZgene resulted in strains with a slightly reduced growth rate, although they were still able to sporulate. The biosynthesis of actinorhodin and, particularly, that of undecylprodigiosin were drastically reduced in the ΔrpoZstrain, suggesting that expression of these secondary metabolite biosynthetic genes is dependent upon the presence of RpoZ in the RNAP complex. Complementation of the ΔrpoZmutant with the wild-typerpoZallele restored both phenotype and antibiotic production. Interestingly, therpoZgene contains a PHO box in its promoter region. DNA binding assays showed that the phosphate response regulator PhoP binds to such a region. Since luciferase reporter studies showed thatrpoZpromoter activity was increased in a ΔphoPbackground, it can be concluded thatrpoZis controlled negatively by PhoP, thus connecting phosphate depletion regulation with antibiotic production and morphological differentiation inStreptomyces.

Download Full-text

Genome-wide analysis of the role of the antibiotic biosynthesis regulator AbsA2 inStreptomyces coelicolorA3(2)

10.1101/361360 ◽

2018 ◽

Author(s):

Richard A. Lewis ◽

Abdul Wahab ◽

Giselda Bucca ◽

Emma E. Laing ◽

Carla Möller-Levet ◽

...

Keyword(s):

Large Scale ◽

Time Course ◽

Streptomyces Coelicolor ◽

Gene Clusters ◽

Biosynthetic Gene Cluster ◽

Negative Influence ◽

Antibiotic Biosynthesis ◽

Biosynthetic Gene

AbstractThe AbsA1-AbsA2 two component signalling system ofStreptomyces coelicolorhas long been known to exert a powerful negative influence on the production of the antibiotics actinorhodin, undecylprodiginine and the Calcium-Dependent Antibiotic (CDA). Here we report the analysis of aΔabsA2deletion strain, which exhibits the classic precocious antibiotic hyper-production phenotype, and its complementation by an N-terminal triple-FLAG-tagged version of AbsA2. The complemented and non-complementedΔabsA2mutant strains were used in large-scale microarray-based time-course experiments to investigate the effect of deletingabsA2on gene expression and to identify thein vivoAbsA2 DNA-binding target sites using ChIP-on chip. We show that in addition to binding to the promoter regions ofredZandactII-orfIVAbsA2 binds to several previously unidentified sites within thecdabiosynthetic gene cluster within and/or upstream ofSCO3215-SCO3216,SCO3217,SCO3229-SCO3230, andSCO3226, and we relate the pattern of AbsA2 binding to the results of the transcriptomic study and antibiotic phenotypic assays. Interestingly, dual ‘biphasic’ ChIP peaks were observed with AbsA2 binding across the regulatory genesactII-orfIVandredZand theabsA2gene itself, while more conventional single promoter-proximal peaks were seen at the CDA biosynthetic genes suggesting a different mechanism of regulation of the former loci. Taken together the results shed light on the complex mechanism of regulation of antibiotic biosynthesis inStreptomyces coelicolorand the important role of AbsA2 in controlling the expression of three antibiotic biosynthetic gene clusters.

Download Full-text

Regulation of the Streptomyces coelicolor Calcium-Dependent Antibiotic by absA, Encoding a Cluster-Linked Two-Component System

Journal of Bacteriology ◽

10.1128/jb.184.3.794-805.2002 ◽

2002 ◽

Vol 184 (3) ◽

pp. 794-805 ◽

Cited By ~ 60

Author(s):

N. Jamie Ryding ◽

Todd B. Anderson ◽

Wendy C. Champness

Keyword(s):

Genomic Sequence ◽

Sequence Data ◽

Streptomyces Coelicolor ◽

Negative Regulation ◽

Negative Regulator ◽

Antibiotic Biosynthesis ◽

Two Component System ◽

Component System ◽

Calcium Dependent ◽

Two Component

ABSTRACT The Streptomyces coelicolor absA two-component system was initially identified through analysis of mutations in the sensor kinase absA1 that caused inhibition of all four antibiotics synthesized by this strain. Previous genetic analysis had suggested that the phosphorylated form of AbsA2 acted as a negative regulator of antibiotic biosynthesis in S. coelicolor (T. B. Anderson, P. Brian, and W. C. Champness, Mol. Microbiol. 39:553–566, 2001). Genomic sequence data subsequently provided by the Sanger Centre (Cambridge, United Kingdom) revealed that absA was located within the gene cluster for production of one of the four antibiotics, calcium-dependent antibiotic (CDA). In this paper we have identified numerous transcriptional start sites within the CDA cluster and have shown that the original antibiotic-negative mutants used to identify absA exhibit a stronger negative regulation of promoters upstream of the proposed CDA biosynthetic genes than of promoters in the clusters responsible for production of actinorhodin and undecylprodigiosin. The same antibiotic-negative mutants also showed an increase in transcription from a promoter divergent to that of absA, upstream of a putative ABC transporter, in addition to an increase in transcription of absA itself. Interestingly, the negative regulation of the biosynthetic transcripts did not appear to be mediated by transcriptional regulation of cdaR (a gene encoding a homolog of the pathway-specific regulators of the act and red clusters) or by any other recognizable transcriptional regulator associated with the cluster. The role of absA in regulating the expression of the diverse antibiotic biosynthesis clusters in the genome is discussed in light of its location in the cda cluster.

Download Full-text

Transmembrane topology of the AbsA1 sensor kinase of Streptomyces coelicolor

Microbiology ◽

10.1099/mic.0.028431-0 ◽

2009 ◽

Vol 155 (6) ◽

pp. 1812-1818 ◽

Cited By ~ 7

Author(s):

Nancy L. McKenzie ◽

Justin R. Nodwell

Keyword(s):

Response Regulator ◽

Streptomyces Coelicolor ◽

Antibiotic Biosynthesis ◽

Sensor Kinase ◽

Response Mechanism ◽

Transmembrane Topology ◽

C Terminus ◽

N Terminus ◽

Sensor Kinases ◽

Insight Into

The sensor kinase AbsA1 (SCO3225) phosphorylates the response regulator AbsA2 (SCO3226) and dephosphorylates AbsA2∼P. The phosphorylated response regulator represses antibiotic biosynthesis operons in Streptomyces coelicolor. AbsA1 was predicted to have an atypical transmembrane topology, and the location of its signal-sensing domain is not readily obvious. To better understand this protein and to gain insight into its signal response mechanism, we determined its transmembrane topology using fusions of absA1 to egfp, which is believed to be the first application of this approach to transmembrane topology in the actinomycetes. Our results are in agreement with the in silico topological predictions and demonstrate that AbsA1 has five transmembrane domains, four near the N terminus and one near the C terminus. Unlike most sensor kinases, the largest extracellular portion of AbsA1 is at the C terminus.

Download Full-text