A type II toxin-antitoxin system is responsible for the cell death at low temperature in Pseudomonas syringae Lz4W lacking RNase R.

RNase R (encoded by the rnr gene) is a highly processive 3' → 5' exoribonuclease essential for the growth of the psychrotrophic bacterium P. syringae Lz4W at low temperature. The cell death of a rnr deletion mutant at low temperature has been previously attributed to the processing defects in 16S rRNA, defective ribosomal assembly and inefficient protein synthesis. We recently showed that RNase R is required to protect P. syringae Lz4W from DNA damage and oxidative stress, independent of its exoribonuclease activity. Here, we show that the processing defect in 16S rRNA does not cause cell death of the rnr mutant of P. syringae at low temperature. Our results demonstrate that the rnr mutant of P. syringae Lz4W, complemented with a RNase R deficient in exoribonuclease function (RNase RD284A) is defective in 16S rRNA processing but can grow at 4 oC. This suggested that the processing defect in ribosomal RNAs is not a cause of the cold sensitivity of the rnr mutant. We further show that the rnr mutant accumulates copies of the indigenous plasmid of P. syringae Lz4W, pLz4W, that bears a type II toxin-antitoxin system (psA-psT). This phenotype was rescued by over-expressing antitoxin psA in the rnr mutant, suggesting that activation of the type II toxin-antitoxin system leads to cold sensitivity of the rnr mutant of P. syringae Lz4W. Here, we report a previously unknown functional relationship between the cold sensitivity of the rnr mutant and a type II toxin-antitoxin system in P. syringae Lz4W.

Exoribonuclease RNase R protects Antarctic Pseudomonas syringae Lz4W from DNA damage and oxidative stress.

IMPORTANCE: Bacterial exoribonucleases play a crucial role in RNA maturation, degradation, quality control, and turnover. In this study, we have uncovered a previously unknown role of 3'-5' exoribonuclease RNase R of Pseudomonas syringae Lz4W in DNA damage and oxidative stress response. Here, we show that neither the exoribonuclease function of RNase R nor its association with the RNA degradosome complex is essential for this function. Interestingly, in P. syringae Lz4W, hydrolytic RNase R exhibits physiological roles similar to phosphorolytic 3'-5' exoribonuclease PNPase of E. coli. Our data suggest that during the course of evolution, mesophilic E. coli and psychrotrophic P. syringae have apparently swapped these exoribonucleases to adapt to their respective environmental growth conditions.

A new role for Escherichia coli Dam DNA methylase in prevention of aberrant chromosomal replication.

The Dam DNA methylase of Escherichia coli is required for methyl-directed mismatch repair, regulation of chromosomal DNA replication initiation from oriC (which is DnaA-dependent), and regulation of gene expression. Here, we show that Dam suppresses aberrant oriC-independent chromosomal replication (also called constitutive stable DNA replication, or cSDR). Dam deficiency conferred cSDR and, in presence of additional mutations (Δtus, rpoB*35) that facilitate retrograde replication fork progression, rescued the lethality of ΔdnaA mutants. The DinG helicase was required for rescue of ΔdnaA inviability during cSDR. Viability of ΔdnaA dam derivatives was dependent on the mismatch repair proteins, since such viability was lost upon introduction of deletions in mutS, mutH or mutL; thus generation of double strand ends (DSEs) by MutHLS action appears to be required for cSDR in the dam mutant. On the other hand, another DSE-generating agent phleomycin was unable to rescue ΔdnaA lethality in dam+ derivatives (mutS+ or ΔmutS), but it could do so in the dam ΔmutS strain. These results point to a second role for Dam deficiency in cSDR. We propose that in Dam-deficient strains, there is an increased likelihood of reverse replication restart (towards oriC) following recombinational repair of DSEs on the chromosome.

Draft Genome Sequence of the Antarctic Psychrophilic Bacterium Pseudomonas syringae Strain Lz4W.

The psychrophilic bacterium Pseudomonas syringae strain Lz4W was isolated from soil samples from Antarctica to decipher the mechanisms of low-temperature adaptation. We report here the 4.982-Mb draft genome sequence of P. syringae Lz4W. This sequence will provide insights into the genomic basis of the psychrophilicity of this bacterium.