Depletion of pre-16S rRNA in starved Escherichia coli cells

Abstract

Specific hybridization assays for intermediates in rRNA synthesis (pre-rRNA) may become useful for monitoring the growth activity of individual microbial species in complex natural systems. This possibility depends upon the assumption that rRNA processing in microbial cells continues after growth and pre-rRNA synthesis cease, resulting in drainage of the pre-rRNA pool. This is not the case in many eukaryotic cells, but less is known about the situation in bacteria. Therefore, we used DNA probes to measure steady-state cellular pre-16S rRNA pools during growth state transitions in Escherichia coli. Pre-16S rRNA became undetectable when cells entered the stationary phase on rich medium and was replenished upon restoration of favorable growth conditions. These fluctuations were of much greater magnitude than concurrent fluctuations in the mature 16S rRNA pool. The extent of pre-16S rRNA depletion depended upon the circumstances limiting growth. It was significantly more pronounced in carbon-energy-starved cells than in nitrogen-starved cells or in cells treated with energy uncouplers. In the presence of the transcriptional inhibitor rifampin, rates of pre-16S rRNA depletion in carbon-energy-starved cells and nitrogen-starved cells were similar, suggesting that the difference between these conditions resides primarily at the level of pre-rRNA synthesis. Chloramphenicol, which inhibits the final steps in rRNA maturation, halted pre-16S rRNA depletion under all conditions. The data show that E. coli cells continue to process pre-rRNA after growth and rrn operon transcription cease, leading to drainage of the pre-rRNA pool. This supports the feasibility of using pre-rRNA-targeted probes to monitor bacterial growth in natural systems, with the caveat that patterns of pre-rRNA depletion vary with the conditions limiting growth.

Relationship between nucleic acid ratios and growth in Listeria monocytogenes

“…Maximum RNA-DNA ratio has been observed at, or close to the end of, exponential growth in E. coli (Davis et al, 1986) and resin acid-degrading bacteria (Muttray & Mohn, 1998), although marine Proteobacteria showed strainspecific responses (Kerkhof & Kemp, 1999). When E. coli was starved and inoculated into fresh medium, a rapid increase in RNA-DNA ratio at the onset of exponential growth was not observed (Cangelosi & Brabant, 1997). During exponential growth, bacteria will tend towards µ max and, in theory, contain maximum cellular concentrations of both rRNA and ribosomes.…”

Monitoring Precursor 16S rRNAs of Acinetobacter spp. in Activated Sludge Wastewater Treatment Systems

“…The RNase III cleavage site is a sequence-independent, secondary-structure-dependent stem produced by intrastrand hybridization between the 5Ј and 3Ј regions of each precursor 16S rRNA molecule. Since no information was available regarding the location of the RNase III cleavage site for precursor 16S rRNA in Acinetobacter spp., probe S-G-Acin-1543-a-A-24 was designed based upon the location of previously reported precursor 16S rRNA probes targeting E. coli (8). In addition, although one species of Acinetobacter is known to have seven copies of the rrn operon (13), no information was available regarding interoperon sequence divergence for Acinetobacter spp.…”

Biomarkers for In Situ Detection of Anaerobic Ammonium-Oxidizing (Anammox) Bacteria

“…Thus, the cellular rRNA content does not reflect the physiological activity of these organisms. For these slowly growing bacteria, the precursor rRNA concentrations are a direct measure of the ribosome turnover rate (Ϸgrowth rate) in the cells (6). To learn more about the in situ activity of anammox organisms, the ISR between the 16S rRNA and 23S rRNA has been targeted with fluorescently labeled oligonucleotide probes.…”