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Expression of ribosome modulation factor (RMF) in Escherichia coli requires ppGpp
1The Institute for Virus Research, Kyoto University, Shogoin-Kawaracho, Sakyo-Ku, Kyoto 606-8507, Japan.
Background:
During the transition from the logarithmic to the stationary phase, 70S ribosomes are dimerized into the 100S form, which has no translational activity. Ribosome Modulation Factor (RMF) is induced during the stationary phase and binds to the 50S ribosomal subunit, which directs the dimerization of 70S ribosomes. Unlike many other genes induced in the stationary phase, rmf transcription is independent of the sigma S. To identify the factors that regulate the growth phase-dependent induction of rmf, mutant strains deficient in global regulators were examined for lacZ expression directed by the rmf promoter.
Results:
Among mutants of defective global regulators, only ppGpp deficiency (relA-spoT double mutant) drastically reduced the level of rmf transcription to less than 10% of that seen in the wild-type. Neither RMF nor 100S ribosomes were detected in this mutant in the stationary phase. rmf transcription correlated well with cellular ppGpp levels during amino acid starvation, IPTG induction of Ptrc-relA455 and in other mutants with artificially increased ppGpp levels. Although the growth rate also correlated inversely with both ppGpp levels and rmf transcription, the observation that the growth rates of the ppGpp-deficient and wild-type strains varied equivalently when grown on different media indicates that the link between rmf transcription and ppGpp levels is not a function of the growth rate.
Conclusions:
ppGpp appears to positively regulate rmf transcription, at least in vivo. Thus, RMF provides a novel negative translational control by facilitating the formation of inactive ribosome dimers (100S) under the stringent circumstances of the stationary phase.
Insights
Guanosine tetraphosphate (ppGpp) positively regulates the transcription of Ribosome Modulation Factor (RMF), which forms inactive 100S ribosome dimers during the stationary phase.
Area of Science:
- Microbiology
- Molecular Biology
- Bacterial Physiology
Background:
- During the transition to the stationary phase, 70S ribosomes dimerize into inactive 100S forms.
- Ribosome Modulation Factor (RMF) induces this dimerization by binding to the 50S ribosomal subunit.
- RMF transcription is independent of sigma S, unlike many other stationary phase genes.
Purpose of the Study:
- To identify factors regulating growth phase-dependent induction of rmf transcription.
- To investigate the role of global regulators in rmf gene expression.
Main Methods:
- Examined lacZ expression directed by the rmf promoter in mutant strains deficient in global regulators.
- Assessed rmf transcription and 100S ribosome formation in ppGpp-deficient mutants.
- Correlated rmf transcription with cellular ppGpp levels under various conditions.
Main Results:
- ppGpp deficiency (relA-spoT double mutant) drastically reduced rmf transcription to less than 10% of wild-type levels.
- RMF and 100S ribosomes were undetectable in ppGpp-deficient mutants during the stationary phase.
- rmf transcription strongly correlated with cellular ppGpp levels, independent of growth rate.
Conclusions:
- ppGpp positively regulates rmf transcription in vivo.
- RMF mediates a novel negative translational control by forming inactive 100S ribosome dimers.
- This mechanism is crucial under the stringent conditions of the stationary phase.
