Development and application of versatile high density microarrays for genome-wide analysis of Streptomyces coelicolor: characterization of the HspR regulon
Bucca, G, Laing, E, Mersinias, V, Allenby, N, Hurd, D, Holdstock, J, Brenner, V, Harrison, M and Smith, CP (2009) Development and application of versatile high density microarrays for genome-wide analysis of Streptomyces coelicolor: characterization of the HspR regulon GENOME BIOL, 10 (1), R5.
development_&_application_BUCCA_09.pdf - Version of Record
Background: DNA microarrays are a key resource for global analysis of genome content, gene expression and the distribution of transcription factor binding sites. We describe the development and application of versatile high density ink-jet in situ-synthesized DNA arrays for the G+C rich bacterium Streptomyces coelicolor. High G+C content DNA probes often perform poorly on arrays, yielding either weak hybridization or non-specific signals. Thus, more than one million 60-mer oligonucleotide probes were experimentally tested for sensitivity and specificity to enable selection of optimal probe sets for the genome microarrays. The heat-shock HspR regulatory system of S. coelicolor, a well-characterized repressor with a small number of known targets, was exploited to test and validate the arrays for use in global chromatin immunoprecipitation-on-chip (ChIP-chip) and gene expression analysis. Results: In addition to confirming dnaK, clpB and lon as in vivo targets of HspR, it was revealed, using a novel ChIPchip data clustering method, that HspR also apparently interacts with ribosomal RNA (rrnD operon) and specific transfer RNA genes (the tRNAGln/tRNAGlu cluster). It is suggested that enhanced synthesis of Glu-tRNAGlu may reflect increased demand for tetrapyrrole biosynthesis following heat-shock. Moreover, it was found that heatshock- induced genes are significantly enriched for Gln/Glu codons relative to the whole genome, a finding that would be consistent with HspR-mediated control of the tRNA species. Conclusions: This study suggests that HspR fulfils a broader, unprecedented role in adaptation to stresses than previously recognized - influencing expression of key components of the translational apparatus in addition to molecular chaperone and protease-encoding genes. It is envisaged that these experimentally optimized arrays will provide a key resource for systems level studies of Streptomyces biology.
|Divisions :||Faculty of Health and Medical Sciences > School of Biosciences and Medicine > Department of Microbial and Cellular Sciences|
|Identification Number :||10.1186/gb-2009-10-1-r5|
|Uncontrolled Keywords :||HEAT-SHOCK RESPONSE, DIFFERENTIALLY EXPRESSED GENES, RIBOSOMAL-RNA SYNTHESIS, ESCHERICHIA-COLI, HELICOBACTER-PYLORI, STRESS-RESPONSE, DNAK OPERON, CHROMATIN IMMUNOPRECIPITATION, MOLECULAR CHAPERONES, NEGATIVE REGULATION|
|Depositing User :||Melanie Hughes|
|Date Deposited :||26 Oct 2010 08:41|
|Last Modified :||23 Sep 2013 18:39|
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