Role of MotA transcription factor in bacteriophage T4 DNA replication

At least two bacteriophage T4 replication origins, ori(uvsY) and ori(34), contain a T4 middle-mode promoter that is necessary for origin function. We wanted to analyze the requirement of these two replication origins for the MotA protein, which is the phage-encoded activator of middle-mode promoters. To ensure the complete absence of MotA protein, we deleted the motA gene from the T4 genome. Unexpectedly, the deletion mutant was not viable unless the MotA protein was provided from a recombinant plasmid. Therefore, MotA is an essential protein for T4 growth. The motA delta mutation reduced the synthesis of several proteins that are encoded by genes with middle-mode promoters, delayed and reduced the synthesis of late proteins, and substantially reduced phage genomic replication. The motA delta mutation also reduced the replication of an ori(uvsY)-containing plasmid and virtually abolished replication of an ori(34)-containing plasmid. The replication defects of the two origins correlated with transcriptional defects: the motA delta mutation modestly reduced transcription from the plasmid-borne ori(uvsY) promoter and strongly reduced transcription from the ori(34) promoter. These results provide strong evidence that MotA protein is normally involved in origin-dependent replication. However, MotA is not required for origin-directed replication as long as transcription can occur from the origin promoter.