A family of anti-σ70 proteins in T4-type phages and bacteria that are similar to AsiA, a transcription inhibitor and co-activator of bacteriophage T4

Anti-sigma(70) factors interact with sigma(70) proteins, the specificity subunits of prokaryotic RNA polymerase. The bacteriophage T4 anti-sigma(70) protein, AsiA, binds tightly to regions 4.1 and 4.2 of the sigma(70) subunit of Escherichia coli RNA polymerase and inhibits transcription from sigma(70) promoters that require recognition of the canonical sigma(70) - 35 DNA sequence. In the presence of the T4 transcription activator MotA, AsiA also functions as a co-activator of transcription from T4 middle promoters, which retain the canonical sigma(70) - 10 consensus sequence but have a MotA box sequence centered at - 30 rather than the sigma(70) - 35 sequence. The E. coli anti-sigma(70) protein Rsd also interacts with region 4.2 of sigma(70) and inhibits transcription from sigma(70) promoters. Our sequence comparisons of T4 AsiA with Rsd, with the, predicted AsiA orthologs of the T4-type phages RB69, 44RR, KVP40, and, - Aeh1, and with AlgQ, a regulator of alginate production in Pseudomonas aeruginosa indicate that these proteins share conserved amino acid residues at positions known to be important for the binding of T4 AsiA to sigma(70) region 4. We show that, like T4 AsiA, Rsd binds to sigma(70) in a native protein gel and, as with T4 AsiA, a L18S substitution in Rsd disrupts this complex. Previous work has assigned sigma(70) amino acid F563, within region 4.1, as a critical determinant for AsiA binding. This residue is also involved in the binding of sigma(70) to the D-flap of core, suggesting that AsiA inhibits transcription by disrupting the interaction between sigma(70) region 4.1 and the beta-flap. We find that as with T4 AsiA, the interaction of KVP40 AsiA, Rsd, or AlgQ with sigma(70) region 4 is diminished by the substitution F563Y. We also demonstrate that like T4 AsiA and Rsd, KVP40 AsiA inhibits transcription from sigma(70)-dependent promoters. We speculate that the phage AsiA orthologs, Rsd, and AlgQ are members of a related family in T4-type phage and bacteria, which interact similarly with primary a factors. In addition, we show that even though a clear MotA ortholog has not been identified in the KVP40 genome and the phage genome appears to lack typical middle promoter sequences, KVP40 AsiA activates transcription from T4 middle promoters in the presence of T4 MotA. We speculate that KVP40 encodes a protein that is dissimilar in sequence, but functionally equivalent, to T4 MotA. Published by Elsevier Ltd.