The control of ribonucleic acid synthesis in bacteria. The synthesis and stability of ribonucleic acid in rifampicin-inhibited cultures of Escherichia coli

Abstract

A study was made of the kinetics of labelling of the stable ribonucleic acids (rRNA+tRNA) and the unstable mRNA fraction in cultures of Escherichia coli M.R.E.600, inhibited by the addition of 0.1g of rifampicin/l. Labelling was carried out by adding either [2-14C]- or [5-3H]-uracil as an exogenous precursor of the cellular nucleic acids. From studies using DNA RNA hybridization, the kinetics of the synthesis and degradation of mRNA was followed in the inhibited cultures. Although a considerable proportion of the mRNA labelled in the presence of rifampicin decayed to non-hybridizable products, about 25% was stabilized beyond the point where protein synthesis had finally ceased. It therefore seems unwise to extrapolate the results of studies on mRNA stability in rifampicin-inhibited cultures to the situation existing in the rate of steady growth, where there appears to be little, if any, stable messenger. The kinetics of labelling of RNA in inhibited cultures indicated that the clapsed time from the addition of rifampicin to the point at which radioactivity no longer enters the total cellular ribonucleic acids is a measure of the time required to polymerize a molecule of rRNA. At 37°C, in culture grown in broth, glucose–salts or lactate salts media, exogenous [2-14C]uracil entered rifampicin-inhibited cells and was incorporated into RNA for 2 3min after the antibiotic was added. Taking this time as that required to polymerize a complete chain of 23S rRNA, the polymerization rate of this fraction in the three media was 25, 22 and 19 nucleotides added/s to the growing chains. Similar experiments in cultures previously inhibited by 0.2g of chloramphenicol/l showed virtually identical behaviour. This confirmed the work of Midgley & Gray (1971), who, by a different approach, showed that the polymerization rate of rRNA in steadily growing and chloramphenicol-inhibited cultures of E. coli at 37°C was essentially constant at about 22 nucleotides added/s. It was thus confirmed that the rate of polymerization of at least the rRNA fraction in E. coli is virtually unaffected by the nature of the growth medium and therefore by bacterial growth rate.