Bacillus cereus ATCC 14579 possesses five RNA helicase-encoding genes overexpressed under cold growth conditions. Out of the five corresponding mutants, only the ⌬cshA, ⌬cshB, and ⌬cshC strains were cold sensitive. Growth of the ⌬cshA strain was also reduced at 30°C but not at 37°C. The cold phenotype was restored with the cshA gene for the ⌬cshA strain and partially for the ⌬cshB strain but not for the ⌬cshC strain, suggesting different functions at low temperature.

In this study, growth rates and lag times of the five RNA helicase-deleted mutants of Bacillus cereus ATCC 14579 were compared to those of the wild-type strain under thermal, oxidative, and pH stresses. Deletion of cshD and cshE had no impact under any of the tested conditions. Deletion of cshA, cshB, and cshC abolished growth at 12°C, confirming previous results. In addition, we found that each RNA helicase had a role in a specific temperature range: deletion of cshA reduced growth at all the tested temperatures up to 45°C, deletion of cshB had impact below 30°C and over 37°C, and deletion of cshC led mainly to a cold-sensitive phenotype. Under oxidative conditions, deletion of cshB and cshC reduced growth rate and increased lag time, while deletion of cshA increased lag time only with H 2 O 2 and reduced growth rate at a high diamide concentration. Growth of the ⌬cshA strain was affected at a basic pH independently of the temperature, while these conditions had a limited effect on ⌬cshB and ⌬cshC strain growth. The RNA helicases CshA, CshB, and CshC could participate in a general adaptation pathway to stressful conditions, with a stronger impact at low temperature and a wider role of CshA.The DEAD-box RNA helicases are encoded by viral, archaeal, eukaryotic, and prokaryotic genomes (9) and play an important role in RNA processing, transport, and degradation and in many other processes involving RNA (4, 19, 26), such as translation or ribosome biogenesis (10,11,22). DEAD-box RNA helicases act as molecular motors that unwind doublestranded RNA, thereby affecting the rearrangement of RNA secondary structures (9, 21). RNA helicases could also be implicated in rearrangement of ribonucleoprotein (RNP) complexes by removing protein from RNA or by the combination of both RNA-unwinding and RNA-annealing activity to promote RNA strand exchange through a potential branch migration (5,13,17,24). Bacterial cells often encounter stressful conditions that tend to decrease the cellular fitness. Consequently, bacteria have to maintain RNA pathway functionalities and control their RNA turnover. Most of the synthesized mRNA is rapidly degraded to allow adaptation to environmental changes (14). RNA helicases could be involved in stress adaptation by maintaining and regulating RNA functions.Studies reporting the involvement of prokaryotic RNA helicases in the adaptation to abiotic stress mainly deal with response to cold, light, and salt conditions (17). The RNA helicase CrhC maintains the photosynthetic capacity of the cyanobacterium Synechocystis. Its expression is regulated by the changes on the redox potential of the electron transport chain caused by variations in light, temperature, and salt concentrations (12). CrhC catalyzes the unwinding of RNA secondary structures but also ensures rearrangements in RNA complexes (5,25). A Bacillus subtilis CshA homolog of Clostridium perfringens is involved in the adaptation to oxidative stress, with the corresponding null mutant strain showing better survival under oxidative stress cond...

Transposon mutagenesis of Bacillus cereus ATCC 14579 yielded cold-sensitive mutants. Mutants of genes encoding enzymes of the central metabolism were affected by cold, but also by other stresses, such as pH or salt, whereas a mutant with transposon insertion in the promoter region of BC0259 gene, encoding a putative DEAD-box RNA helicase displaying homology with Escherichia coli CsdA and Bacillus subtilis CshA RNA helicases, was only cold-sensitive. Expression of the BC0259 gene at 10 degrees C is reduced in the mutant. Analysis of the 5' untranslated region revealed the transcriptional start and putative cold shock-responsive elements. The role of this RNA helicase in the cold-adaptive response of B. cereus is discussed.