Control of the Synthesis of T4 Phage Deoxynucleotide Kinase Messenger Ribonucleic Acid in Vivo

Abstract

Abstract The regulation of the synthesis of messenger RNA of T4 phage-specific deoxynucleotide kinase has been studied in vivo by assay of mRNA for its capacity to direct the synthesis of deoxynucleotide kinase in vitro. The deoxynucleotide kinase mRNA first appears at 2 min after infection and increases in amount until 8 min. Thereafter the messenger activity decreases. The appearance of deoxynucleotide kinase mRNA is blocked when chloramphenicol is added before or immediately after infection. When added at 8 min the normal levels of kinase mRNA were observed at times later in the infection period. However, the addition of chloramphenicol at 3 min after infection results in the production of an exaggerated amount of deoxynucleotide kinase mRNA, which keeps increasing until at least 30 min after infection. The high level of mRNA formed in the presence of chloramphenicol could reflect the absence of an agent or agents required for the termination of the synthesis of this mRNA. This conclusion was reached from experiments with the inhibitor of RNA synthesis, rifampicin. When added to cultures at a time late in the infection period, rifampicin caused an immediate drop in the level of kinase mRNA. At this time the level of this mRNA is therefore a balance between synthesis and degradation. When similar experiments were carried out to study the level of mRNA for α-glucosyltransferase, cultures receiving chloramphenicol at 3 min after infection and rifampicin at 11 and 17 min did not show a drop in the level of mRNA for this enzyme. There is thus a difference in the stability of the two types of mRNA in regard to their degradation by bacterial nucleases when the infection was carried out with chloramphenicol added at 3 min. In the absence of chloramphenicol both kinds of mRNA underwent degradation. The overproduction and the delay of shutoff of the synthesis of deoxynucleotide kinase mRNA are also seen when nonpermissive Escherichia coli K-12 is infected with T4 DO mutants or when DNA synthesis is inhibited by fluorodeoxyuridine. The infection with an MD mutant does not result in the same effect as that observed with the DO mutants. We favor the hypothesis that both new T4 protein (or proteins) and continuous T4 DNA replication are necessary for the proper termination of synthesis of delayed-early T4 RNA.