| Autor: |
Wilson, Stuart E., Ingham, Colin J., Hunter, Iain S., Smith, Margaret C. M. |
| Předmět: |
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| Zdroj: |
Molecular Microbiology; Apr1995, Vol. 16 Issue 1, p131-143, 13p, 6 Diagrams, 1 Chart |
| Abstrakt: |
The repressor gene, c, is required for maintenance of lysogeny in the Streptomyces phage φC31. The c gene expresses three in-frame N-terminally different protein isoforms at least one of which is thought to bind to a 17 bp highly conserved inverted repeat (CIR) sequence found at 18 (or more) loci throughout the φC31 genome. Here we present evidence that one of these loci, CIR6, and its interaction with the products of the repressor gene are critical in the control of the lytic pathway in φC31. To the right of CIR6, according to the standard map of φC31, an 'immediate-early' promoter, ap1, was discovered after insertion of a fragment containing CIR6 upstream of a promoterless kanamycin-resistance gene, aphll, to form pCIA2. pCIA2 conferred kanamycin resistance upon Streptomyces coelicolor A3(2) but not upon a φC31 lysogen of S. coelicolor. Operator-constitutive (Oc) mutants of pCIA2 were isolated and the mutations lay in CIR6, i.e. C(R6:G14T and CIR6:C2A. Primer extension analysis of RNA prepared from an induced, temperature-sensitive lysogen of S. coelicolor localized a mRNA 5′ endpoint 21 bp to the right of CIR6. The importance of the ap1/CIR6 region in the regulation of lytic growth was demonstrated by the analysis of a virulent mutant, φC31 vir1, capable of forming plaques on an S. coelicolor φC31 lysogen. φC31 vir1 contained a DNA inversion with the breakpoints lying within the integrase gene (which lies approximately 7kbp to the right of CIR6) and in the essential early region between CIR6 and the -- 10 sequence for ap1. The separation of ap1 from its operator was thought to be the basis for the virulent phenotype in φC31 vir1. Band-shift assays and DNase I footprinting experiments using purified 42 kDa repressor isoform confirmed that CIRs 5 and 6 were indeed the targets for binding of th... [ABSTRACT FROM AUTHOR] |
| Databáze: |
Complementary Index |
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