ABSTRACT
Escherichia coli phage phiEco32 encodes two proteins that bind to host RNA polymerase (RNAP): gp79, a novel protein, and gp36, a distant homolog of σ(70) family proteins. Here, we investigated the temporal pattern of phiEco32 and host gene expression during infection. Host transcription shutoff and three distinct bacteriophage temporal gene classes (early, middle, and late) were revealed. A combination of bioinformatic and biochemical approaches allowed identification of phage promoters recognized by a host RNAP holoenzyme containing the σ(70) factor. These promoters are located upstream of early phage genes. A combination of macroarray data, primer extension, and in vitro transcription analyses allowed identification of six promoters recognized by an RNAP holoenzyme containing gp36. These promoters are characterized by a single-consensus element tAATGTAtA and are located upstream of the middle and late phage genes. Curiously, gp79, an inhibitor of host and early phage transcription by σ(70) holoenzyme, activated transcription by the gp36 holoenzyme in vitro.
Subject(s)
Coliphages/genetics , Escherichia coli/virology , Gene Expression Regulation, Viral , Base Sequence , Computational Biology , DNA-Directed RNA Polymerases/metabolism , Molecular Sequence Data , Promoter Regions, Genetic , Sigma Factor/metabolism , Transcription, Genetic , Transcriptional ActivationABSTRACT
A novel bacteriophage infecting Escherichia coli was isolated during a large-scale screen for bacteriophages that may be used for therapy of mastitis in cattle. The 77,554-bp genome of the bacteriophage, named phiEco32, was sequenced and annotated, and its virions were characterized by electron microscopy and proteomics. Two phiEco32-encoded proteins that interact with host RNA polymerase were identified. One of them is an ECF family sigma factor that may be responsible for transcription of some viral genes. Another RNA polymerase-binding protein is a novel transcription inhibitor whose mechanism of action remains to be defined.
