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A Novel Transcriptional Activator, tubX , Is Required for the Stability of Bacillus sphaericus Mosquitocidal Plasmid pBsph
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| Zeitschriftentitel: | Journal of Bacteriology |
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| Personen und Körperschaften: | , , , , , |
| In: | Journal of Bacteriology, 196, 2014, 24, S. 4304-4314 |
| Format: | E-Article |
| Sprache: | Englisch |
| veröffentlicht: |
American Society for Microbiology
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| Schlagwörter: |
| Zusammenfassung: | <jats:title>ABSTRACT</jats:title> <jats:p> Stable maintenance of the low-copy-number plasmid pBsph in <jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">Bacillus sphaericus</jats:named-content> requires a partitioning ( <jats:italic>par</jats:italic> ) system that consists of a filament-forming protein, <jats:named-content xmlns:xlink="http://www.w3.org/1999/xlink" content-type="genus-species" xlink:type="simple">B. sphaericus</jats:named-content> TubZ (TubZ-Bs); a centromere-binding protein, TubR-Bs; and a centromere-like DNA site, <jats:italic>tubC</jats:italic> , composed of three blocks (I, II, and III) of 12-bp degenerate repeats. Previous studies have shown that mini-pBsph replicons encoding the TubZ system are segregationally highly unstable, whereas the native pBsph is stably maintained. However, the mechanism underlying the stability discrepancy between pBsph and its minireplicon is poorly understood. Here <jats:italic>orf187</jats:italic> (encoding TubX), a gene downstream of <jats:italic>tubZ</jats:italic> -Bs, was found to play a role in plasmid stabilization. Null mutation or overexpression of <jats:italic>tubX</jats:italic> resulted in a defect in pBsph stability and a significant decrease in the level of <jats:italic>tubRZ</jats:italic> -Bs expression, and the TubX-null phenotype was suppressed by ectopic expression of a wild-type copy of <jats:italic>tubX</jats:italic> and additional <jats:italic>tubRZ</jats:italic> -Bs. An electrophoresis mobility shift assay (EMSA) and a DNase I footprinting assay revealed that the TubX protein bound directly to five 8-bp degenerate repeats located in the <jats:italic>par</jats:italic> promoter region and that TubX competed with TubR-Bs for binding to the <jats:italic>par</jats:italic> promoter. Further studies demonstrated that TubX significantly stimulated the transcription of the <jats:italic>par</jats:italic> operon in the absence of <jats:italic>tubR</jats:italic> -Bs, and a higher level of gene activation was observed when <jats:italic>tubR</jats:italic> -Bs was present. These results suggested that TubX positively regulates <jats:italic>tubRZ</jats:italic> -Bs transcription by interfering with TubR-Bs-mediated repression and binding directly to the <jats:italic>tubRZ</jats:italic> -Bs promoter region. </jats:p> |
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| Umfang: | 4304-4314 |
| ISSN: |
0021-9193
1098-5530 |
| DOI: | 10.1128/jb.01855-14 |