Phage T4 early promoters are resistant to inhibition by the anti-sigma factor AsiA.

Abstract : Phage T4 early promoters are transcribed in vivo and in vitro by the Escherichia coli RNA polymerase holoenzyme Esigma(70). We studied in vitro the effects of the T4 anti-sigma(70) factor AsiA on the activity of several T4 early promoters. In single-round transcription, promoters motB, denV, mrh.2, motA wild type and UP element-deleted motA are strongly resistant to inhibition by AsiA. The alpha-C-terminal domain of Esigma(70) is crucial to this resistance. DNase I footprinting of Esigma(70) and Esigma(70)AsiA on motA and mrh.2 shows extended contacts between the holoenzyme with or without AsiA and upstream regions of these promoters. A TG --> TC mutation of the extended -10 motif in the motA UP element-deleted promoter strongly increases susceptibility to inhibition by AsiA, but has no effect on the motA wild-type promoter: either the UP element or the extended -10 site confers resistance to AsiA. Potassium permanganate reactivity shows that the two structure elements are not equivalent: with AsiA, the motA UP element-deleted promoter opens more slowly whereas the motA TC promoter opens like the wild type. Changes in UV laser photoreactivity at position +4 on variants of motA reveal an analogous distinction in the roles of the extended -10 and UP promoter elements.
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Gilbert Orsini, Sébastien Igonet, Carole Pène, Bianca Sclavi, Malcolm Buckle, et al.. Phage T4 early promoters are resistant to inhibition by the anti-sigma factor AsiA.. Molecular Microbiology, Wiley, 2004, 52 (4), pp.1013-28. ⟨10.1111/j.1365-2958.2004.04038.x⟩. ⟨hal-00277452⟩



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