The transcription termination factor Rho is required for oxidative stress survival in Caulobacter crescentus

Abstract

Summary A transposon Tn5 mutagenesis library was generated from Caulobacter crescentus strain NA1000, and clones with deficiency in survival in a high concentration of NaCl were selected. One of these clones, 37G10, has the Tn5 integrated within the coding region of the transcription termination factor Rho. Analysis of this mutant phenotype showed that the cells are motile and present a normal cell cycle, but have a longer generation time. This strain is sensitive to acidic pH, to the presence of different salts and to heat shock, but it responds well to UV light and alkaline pH. The most striking phenotype of the rho mutant is that it is extremely sensitive to oxidative stress, in both exponential and stationary phases. Experiments using a transcriptional fusion of the rho promoter region to the lacZ gene showed that rho gene expression varies during the cell cycle, showing very low expression levels at the swarmer cell stage and presenting maximum levels in early predivisional cells. Transcription of the rho gene is increased in the rho mutant strain, which is indicative of an autoregulatory circuit, and there is a small variation in the cell cycle pattern of expression. Several peptides have their synthesis altered in the mutant strain, as analysed by two‐dimensional gel electrophoresis, most of which show a reduction in expression. These results indicate that the Rho factor is essential for an efficient response to certain stresses in Caulobacter.

Torn apart and reunited: impact of a heterotroph on the transcriptome of Prochlorococcus

“…Changes in the cellular redox state during co-culture may have been part of the signal sensed by the cell to elicit these responses, either directly transcription factors as described above, or through post-transcriptional regulation. Indeed, the abundance of Rho transcripts, a gene that has an important role in regulating transcriptional termination and contributes to oxidative stress survival in other bacteria (Italiani et al, 2002;Kawamura et al, 2005) decreased after 24 h of co-culture.…”

Prediction of Transcriptional Terminators in Bacillus subtilis and Related Species

“…However, the relative importance of Rhodependent termination varies between bacteria. In the proteobacteria E. coli and Rhodobacter sphaeroides, as well as the actinobacterium Micrococcus luteus, Rho is essential [10][11][12]; in the proteobacterium Caulobacter crescentus, Rho is required for oxidative stress survival [13]. Also, an analysis of the average RNA folding energy near stop codons suggested that no stem-loops are formed in the Firmicutes Mycoplasma genitalium and M. pneumoniae, the actinobacterium Mycobacterium pneumoniae, the cyanobacterium Synechocystis sp.…”

Isolation and characterization of Caulobacter mutants impaired in adaptation to stationary phase

“…The cells were tested for oxidative, saline, acidic, heat and UV exposure (Fig. 2), at conditions in which the wild type cells were not affected (12). As a second control, a mutant strain with the Tn5 inserted into the rsaF gene that encodes an outer membrane protein (OMP58) was tested for all stresses and responded like the wild type, indicating that the sensitivity to stress is not due to the presence of the Tn5 (not shown).…”

“…Tn5 (Kan) transposon was inserted into C. crescentus strain NA1000 by conjugation with an E. coli S17-1 carrying plasmid pSUP2021 (25) as described (12). Screeening was performed as described previously (28), with the following modifications: the cells were grown in 96-well plates containing 0.3 ml of PYE/Kan medium at 30ºC for 14 days with agitation, followed by plating in PYE/Kan.…”

“…The determination of the Tn5 insertion sites was performed by reverse PCR as described (12). The PCR conditions were: 5 minutes 95ºC, 30 cycles of 1 minute 95ºC, 1 minute 42ºC or 52ºC and 1 minute 72ºC, and a final cycle of 7 minutes at 72ºC.…”