Development and application of versatile high density microarrays for genome-wide analysis of Streptomyces coelicolor: characterization of the HspR regulon

Bucca, Giselda; Laing, Emma; Mersinias, Vassilis; Allenby, Nicholas E. E.; Hurd, Douglas; Holdstock, Jolyon; Brenner, Volker; Harrison, Marcus; Smith, Colin P.

doi:10.1186/gb-2009-10-1-r5

Cited by 37 publications

(45 citation statements)

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“…Therefore, the GlxR-binding sites located far from the coding region could not be detected by the microarray, as has been described by others (27,52). False negatives may also occur in ChIP-chip experiments due to sequestration of the transcription factor in nucleoprotein complexes, rendering them inaccessible to the specific test antibody used for the IP reaction (12,30). Of the binding sites not detected by ChIP-chip analyses in this study, 21 have been shown to bind to GlxR in EMSA (42,43).…”

Section: Vol 193 2011mentioning

confidence: 92%

“…In addition, the cyaB mutant is able to grow on any carbon source much better than a glxR mutant (15,59), implying that GlxR may be physiologically functional even in the cyaB mutant background. Chromatin immunoprecipitation in conjunction with microarray (ChIP-chip) analysis has been used to identify in vivo target sites and physiological states of bacterial transcriptional regulators in recent years (12,17,27,30,31,52,60,66). The regions bound by the transcriptional regulator are enriched by ChIP, and the degrees of the enrichment reflect the ability of the regulator to bind to the regions in the cells under the conditions tested.…”

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confidence: 99%

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Genome-Wide Identification of In Vivo Binding Sites of GlxR, a Cyclic AMP Receptor Protein-Type Regulator in Corynebacterium glutamicum

et al. 2011

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Corynebacterium glutamicum GlxR is a cyclic AMP (cAMP) receptor protein-type regulator. Although over 200 GlxR-binding sites in the C. glutamicum genome are predicted in silico, studies on the physiological function of GlxR have been hindered by the severe growth defects of a glxR mutant. This study identified the GlxR regulon by chromatin immunoprecipitation in conjunction with microarray (ChIP-chip) analyses. In total, 209 regions were detected as in vivo GlxR-binding sites. In vitro binding assays and promoter-reporter assays demonstrated that GlxR directly activates expression of genes for aerobic respiration, ATP synthesis, and glycolysis and that it is required for expression of genes for cell separation and mechanosensitive channels. GlxR also directly represses a citrate uptake gene in the presence of citrate. Moreover, ChIP-chip analyses showed that GlxR was still able to interact with its target sites in a mutant with a deletion of cyaB, the sole adenylate cyclase gene in the genome, even though binding affinity was markedly decreased. Thus, GlxR is physiologically functional at the relatively low cAMP levels in the cyaB mutant, allowing the cyaB mutant to grow much better than the glxR mutant.

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Section: Vol 193 2011mentioning

confidence: 92%

mentioning

confidence: 99%

Genome-Wide Identification of In Vivo Binding Sites of GlxR, a Cyclic AMP Receptor Protein-Type Regulator in Corynebacterium glutamicum

et al. 2011

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“…Before use, the RNA samples were checked for integrity on an Agilent 2100 Bioanalyzer (Agilent Technologies), and lack of DNA was checked by reverse transcription-PCR (RT-PCR) without reverse transcriptase. cDNA preparation, labeling, and hybridization was performed as previously described (41,42). For the microarray analysis, 4ϫ44K whole-genome Streptomyces arrays (Agilent Technologies) were used (42).…”

Section: Methodsmentioning

confidence: 99%

“…cDNA preparation, labeling, and hybridization was performed as previously described (41,42). For the microarray analysis, 4ϫ44K whole-genome Streptomyces arrays (Agilent Technologies) were used (42). The filtered data sets were analyzed using rank product analysis (43) via the Rank-Prod package in R (version 2.5.0) (44).…”

Section: Methodsmentioning

confidence: 99%

The ROK Family Regulator Rok7B7 Pleiotropically Affects Xylose Utilization, Carbon Catabolite Repression, and Antibiotic Production in Streptomyces coelicolor

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Gubbens

Bucca

et al. 2013

Journal of Bacteriology

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cMembers of the ROK family of proteins are mostly transcriptional regulators and kinases that generally relate to the control of primary metabolism, whereby its member glucose kinase acts as the central control protein in carbon control in Streptomyces. Here, we show that deletion of SCO6008 (rok7B7) strongly affects carbon catabolite repression (CCR), growth, and antibiotic production in Streptomyces coelicolor. Deletion of SCO7543 also affected antibiotic production, while no major changes were observed after deletion of the rok family genes SCO0794, SCO1060, SCO2846, SCO6566, or SCO6600. Global expression profiling of the rok7B7 mutant by proteomics and microarray analysis revealed strong upregulation of the xylose transporter operon xylFGH, which lies immediately downstream of rok7B7, consistent with the improved growth and delayed development of the mutant on xylose. The enhanced CCR, which was especially obvious on rich or xylose-containing media, correlated with elevated expression of glucose kinase and of the glucose transporter GlcP. In liquid-grown cultures, expression of the biosynthetic enzymes for production of prodigionines, siderophores, and calcium-dependent antibiotic (CDA) was enhanced in the mutant, and overproduction of prodigionines was corroborated by matrixassisted laser desorption ionization-time-of-flight analysis. These data present Rok7B7 as a pleiotropic regulator of growth, CCR, and antibiotic production in Streptomyces.

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“…The feature extraction output files were imported into R (software programming language) (version 2.5.0; http://www.R-project.org) and normalized using the LIMMA package (18); global median within-array normalization followed by "scale" across-array normalization was applied to the log 2 [cDNA/gDNA] ratios of the expression arrays. Flagging of poor-quality spots was performed (19), and probes were filtered out of the data set if they did not yield a good-quality spot across gene expression data sets. Probes targeting the coding regions of a gene were averaged such that each annotated protein-encoding gene of S. coelicolor was represented by a single value.…”

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confidence: 99%

AterDDomain-Encoding Gene (SCO2368) Is Involved in Calcium Homeostasis and Participates in Calcium Regulation of a DosR-Like Regulon in Streptomyces coelicolor

et al. 2015

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bAlthough Streptomyces coelicolor is not resistant to tellurite, it possesses several TerD domain-encoding (tdd) genes of unknown function. To elucidate the function of tdd8, the transcriptomes of S. coelicolor strain M145 and of a tdd8 deletion mutant derivative (the ⌬tdd8 strain) were compared. Several orthologs of Mycobacterium tuberculosis genes involved in dormancy survival were upregulated in the deletion mutant at the visual onset of prodiginine production. These genes are organized in a putative redox stress response cluster comprising two large loci. A binding motif similar to the dormancy survival regulator (DosR) binding site of M. tuberculosis has been identified in the upstream sequences of most genes in these loci. A predicted role for these genes in the redox stress response is supported by the low NAD ؉ /NADH ratio in the ⌬tdd8 strain. This S. coelicolor gene cluster was shown to be induced by hypoxia and NO stress. While the tdd8 deletion mutant (the ⌬tdd8 strain) was unable to maintain calcium homeostasis in a calcium-depleted medium, the addition of Ca 2؉ in ⌬tdd8 culture medium reduced the expression of several genes of the redox stress response cluster. The results shown in this work are consistent with Tdd8 playing a significant role in calcium homeostasis and redox stress adaptation.

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Development and application of versatile high density microarrays for genome-wide analysis of Streptomyces coelicolor: characterization of the HspR regulon

Abstract: Development of high-density microarrays for global analysis of gene expression and transcription factor binding in Streptomyces coelicolor suggests a novel role for HspR in stress adaptation.

Cited by 37 publications

References 65 publications

Genome-Wide Identification of In Vivo Binding Sites of GlxR, a Cyclic AMP Receptor Protein-Type Regulator in Corynebacterium glutamicum

Genome-Wide Identification of In Vivo Binding Sites of GlxR, a Cyclic AMP Receptor Protein-Type Regulator in Corynebacterium glutamicum

The ROK Family Regulator Rok7B7 Pleiotropically Affects Xylose Utilization, Carbon Catabolite Repression, and Antibiotic Production in Streptomyces coelicolor

AterDDomain-Encoding Gene (SCO2368) Is Involved in Calcium Homeostasis and Participates in Calcium Regulation of a DosR-Like Regulon in Streptomyces coelicolor

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