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Regulation of antibiotic production in Streptomyces coelicolor: Phenotypic and transcriptomic analysis of AbsA mutants.

Wahab, Adbul. (2007) Regulation of antibiotic production in Streptomyces coelicolor: Phenotypic and transcriptomic analysis of AbsA mutants. Doctoral thesis, University of Surrey (United Kingdom)..

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The AbsA two-component system is a negative regulator of antibiotic biosynthesis in Streptomyces coelicolor. To gain further insight into the role of the AbsA system different in-frame deletion mutations were made in the absA1A2 operon in S. coelicolor MT1110, MT1110DeltacdaR and S. lividans 1326. Phenotypic analysis of DeltaabsA1A2 and DeltaabsA2 in the MT1110 derivatives showed that CDA was produced early on Oxoid nutrient agar relative to the parent strain. The absA mutants in S. lividans 1326 (both DeltaabsA1 and DeltaabsA1A2) did not show early CDA production. On solid R2YE both MT1110DeltaabsA1A2 and MT1110DeltaabsA2 produced undecylprodigiosin and actinorhodin earlier. In liquid R2YE medium MT1110DeltaabsA1A2 produced undecylprodigiosin and actinorhodin earlier compared to MT1110 wild-type. RNA was extracted from surface-grown cultures of MT1110 and its DeltaabsA1A2 derivative on R2YE agar at different stages of growth. Cy3-labelled cDNA and Cy5-labelled genomic DNA (gDNA) were hybridised on in-house printed 50-mer oligo arrays representing all S. coelicolor open reading frames. Differentially expressed genes were identified by the 'Rank Product' method. Transcriptomic analysis revealed that many genes encoding enzymes of central metabolism were up-regulated in MT1110DeltaabsA1A2 compared to MT1110 wild-type. Three genes, SCO4089, SCO3829 and SCO1270 encoding valine dehydrogenase, dihydrolipoamide acyltransferase component E2 and the alpha-subunit of pyruvate dehydrogenase complex respectively, were found to be more highly expressed in MT1110DeltaabsA1A2. These genes encode enzymes whose reactions produce possibly precursors for CDA, Act and Red biosynthesis. The gene encoding pyruvate kinase 2 (SCO5423) showed increased expression in the mutant at and after 24 h of growth. Genes encoding subunits of ATP synthases showed increased expression in the mutant that indicates that an increase oxidative metabolism is occurring in MT1110DeltaabsA1A2. A gene, SCO6195, encoding a stationary phase-active enzyme, acyl-coenzyme A synthetase was induced in MT1110DeltaabsA1A2 at 23 and 24 h of cultivation. Two genes in the cda cluster were found to be repressed in mutant: SCO3224 and SCO3229, encoding an ABC transporter ATP-binding protein and 4-hydroxymandelate synthase, respectively. The gene encoding AfsS (SCO4425) was found to be up-regulated in the DeltaabsA1A2 mutant at the late phase of the growth. Four genes encoding enzymes putatively involved in the pentose phosphate pathway were found to be up-regulated in MT1110DeltaabsA1A2 at certain time points. Genes encoding ribosomal protein showed a more relaxed growth phase dependent pattern of expression in the mutant while one r- protein encoding gene, rpsK (SCO4728), showed higher expression in the mutant except at stationary phase. It is deduced from the global gene expression analysis that the possible role of the AbsA system in S. coelicolor is to regulate precursor supply for antibiotic production rather than to directly regulate the genes of the respective antibiotic biosynthetic clusters.

Item Type: Thesis (Doctoral)
Divisions : Theses
Authors :
Wahab, Adbul.
Date : 2007
Contributors :
Depositing User : EPrints Services
Date Deposited : 09 Nov 2017 12:14
Last Modified : 15 Mar 2018 20:24

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