Role of anionic phospholipids in the adaptation of Bacillus subtilis to high salinity

Abstract

The importance of the content of anionic phospholipids [cardiolipin (CL) and phosphatidylglycerol (PG)] in the osmotic adaptation and in the membrane structure of Bacillus subtilis cultures was investigated. Insertion mutations in the three putative cardiolipin synthase genes (ywiE, ywnE and ywjE) were obtained. Only the ywnE mutation resulted in a complete deficiency in cardiolipin and thus corresponds to a true clsA gene. The osmotolerance of a clsA mutant was impaired: although at NaCl concentrations lower than 1?2 M the growth curves were similar to those of its wild-type control, at 1?5 M NaCl (LBN medium) the lag period increased and the maximal optical density reached was lower. The membrane of the clsA mutant strain showed an increased PG content, at both exponential and stationary phase, but no trace of CL in either LB or LBN medium. As well as the deficiency in CL synthesis, the clsA mutant showed other differences in lipid and fatty acids content compared to the wild-type, suggesting a cross-regulation in membrane lipid pathways, crucial for the maintenance of membrane functionality and integrity. The biophysical characteristics of membranes and large unilamellar vesicles from the wild-type and clsA mutant strains were studied by Laurdan's steady-state fluorescence spectroscopy. At physiological temperature, the clsA mutant showed a decreased lateral lipid packing in the protein-free vesicles and isolated membranes compared with the wild-type strain. Interestingly, the lateral lipid packing of the membranes of both the wild-type and clsA mutant strains increased when they were grown in LBN. In a conditional IPTG-controlled pgsA mutant, unable to synthesize PG and CL in the absence of IPTG, the osmoresistance of the cultures correlated with their content of anionic phospholipids. The transcriptional activity of the clsA and pgsA genes was similar and increased twofold upon entry to stationary phase or under osmotic upshift. Overall, these results support the involvement of the anionic phospholipids in the growth of B. subtilis in media containing elevated NaCl concentrations.

Thematic Review Series: Glycerolipids. Cardiolipin synthesis for the assembly of bacterial and mitochondrial membranes

“…The increase in cardiolipin formation seems to be driven by higher expression of cardiolipin synthase (23,24). Recent studies confirmed that bacteria with cardiolipin synthase deficiency are more vulnerable to osmotic stress and organic solvents (96,97). The mechanism by which cardiolipin stabilizes bacterial membranes is not known, but an interesting observation was made recently, namely that cardiolipin localizes to the polar and septal regions of the cytoplasmic membrane (98)(99)(100).…”

A Comprehensive Proteomics and Transcriptomics Analysis ofBacillus subtilisSalt Stress Adaptation

“…In our data, transcription of the clsA gene as well as the ywjE gene is only marginally affected by salt stress, but the B -controlled ywiE gene is induced more than 8-fold. Nevertheless, ywiE and ywjE are not essential during hyperosmotic conditions, whereas clsA is required for increased osmotolerance in B. subtilis (28,40). A third distinct biochemical change in the membrane composition is the increase in unsaturated fatty acids, which is catalyzed by a membrane-located ⌬ 5 -desaturase encoded by the cold-inducible des gene (1).…”

“…Both the W and M regulons are typically induced by cell envelope stress, and M is essential for growth and cellular survival in high-salt concentrations (29,51). Severe salt stress has pleiotropic effects on the physiology of B. subtilis: the composition of the cytoplasmic membrane is altered (40,41), cell wall properties are adjusted (42), and swarming capability of the cells is largely impaired (57).…”

Adaptation beyond the Stress Response: Cell Structure Dynamics and Population Heterogeneity in Staphylococcus aureus

“…CL can stabilize a membrane against changes of osmotic stress when included in liposomes (60). Recently, CL was demonstrated to be important for E. coli and B. subtilis to grow at high salinity (47,72). In contrast, S. aureus does not require CL to grow in high salinity medium (Tsai et al, submitted).…”