Transcriptome and membrane fatty acid analyses reveal different strategies for responding to permeating and non-permeating solutes in the bacterium Sphingomonas wittichii

Abstract

BackgroundSphingomonas wittichii strain RW1 can completely oxidize dibenzo-p-dioxins and dibenzofurans, which are persistent contaminants of soils and sediments. For successful application in soil bioremediation systems, strain RW1 must cope with fluctuations in water availability, or water potential. Thus far, however, little is known about the adaptive strategies used by Sphingomonas bacteria to respond to changes in water potential. To improve our understanding, strain RW1 was perturbed with either the cell-permeating solute sodium chloride or the non-permeating solute polyethylene glycol with a molecular weight of 8000 (PEG8000). These solutes are assumed to simulate the solute and matric components of the total water potential, respectively. The responses to these perturbations were then assessed and compared using a combination of growth assays, transcriptome profiling, and membrane fatty acid analyses.ResultsUnder conditions producing a similar decrease in water potential but without effect on growth rate, there was only a limited shared response to perturbation with sodium chloride or PEG8000. This shared response included the increased expression of genes involved with trehalose and exopolysaccharide biosynthesis and the reduced expression of genes involved with flagella biosynthesis. Mostly, the responses to perturbation with sodium chloride or PEG8000 were very different. Only sodium chloride triggered the increased expression of two ECF-type RNA polymerase sigma factors and the differential expression of many genes involved with outer membrane and amino acid metabolism. In contrast, only PEG8000 triggered the increased expression of a heat shock-type RNA polymerase sigma factor along with many genes involved with protein turnover and repair. Membrane fatty acid analyses further corroborated these differences. The degree of saturation of membrane fatty acids increased after perturbation with sodium chloride but had the opposite effect and decreased after perturbation with PEG8000.ConclusionsA combination of growth assays, transcriptome profiling, and membrane fatty acid analyses revealed that permeating and non-permeating solutes trigger different adaptive responses in strain RW1, suggesting these solutes affect cells in fundamentally different ways. Future work is now needed that connects these responses with the responses observed in more realistic scenarios of soil desiccation.

Exposure to Solute Stress Affects Genome-Wide Expression but Not the Polycyclic Aromatic Hydrocarbon-Degrading Activity of Sphingomonas sp. Strain LH128 in Biofilms

“…An increase in the relative abundance of saturated fatty acids has been proposed to reduce cell membrane fluidity and permeability and, as such, decrease the rate of water loss under osmotic stress (44). This has been shown in S. wittichii RW1 (38), where the ratio was increased by ϳ1.3-fold compared to an ϳ1.8-fold increase in LH128. The apparent lack of response of membrane fatty acid saturation to the acute solute stress could be explained by the fact that changes in membrane fatty acid saturation requires cell growth (26) and that the majority of the biofilm cells did not have time to grow during the 4-h exposure, with the doubling time of LH128 on phenanthrene being 11 h (P. Bruegelmans and D. Springael, unpublished results).…”

“…We have indications for phenanthrene inducible expression of at least phnA1f involved in initial aromatic ring hydroxylation (K. Wouters and D. Springael, unpublished results). Also, in salicylate-grown cells of S. wittichii RW1, the transcription of genes involved in aromatic degradation was not affected by immediate and long-term exposure to NaCl (38).…”

“…Glutamate is an important osmoregulatory amino acid in bacteria exposed to solute stress (13) and the expression of its biosynthesis genes upon solute exposure was also reported in Sinorhizobium meliloti (16). The importance of ectoine as an osmoprotectant response to solute stress was shown for S. meliloti and Desulfovibrio vulgaris (25,34), while biosynthesis of trehalose as a response to NaCl stress was also suggested in S. wittichii RW1 (38). The role of import of compatible solutes is further accentuated by the increased expression of genes encoding proteins that show homology to permeases and conductance mechanosensitive channel (MSC).…”

“…For this purpose, strain LH128 was grown in continuous flow chambers that contain solid phenanthrene as a sole carbon source and that allow easy recovery of biofilm cells for transcriptomic and physiological analysis. As in other studies addressing the bacterial response to solute stress, NaCl was used as the osmolyte (9,38). Both acute and chronic solute stress was invoked to assess differences in short-term and long-term responses.…”

Physiology and transcriptome of the polycyclic aromatic hydrocarbon-degrading Sphingomonas sp. LH128 after long-term starvation

“…As a physiological adaptation, LH128 cells appear to increase the degree of saturation of membrane fatty acids. An increase in the degree of phospholipid fatty acid saturation reduces cell membrane fluidity and permeability and has been previously reported in a number of bacteria as a response to the presence of solutes or toxic hydrocarbons and to a shift in temperature (Heipieper & de Bont, 1994;Heipieper et al, 2003;Johnson et al, 2011;Kieft et al, 1997;Mrozik et al, 2004;Suutari & Laakso, 1994), but has not yet been related to nutrient starvation. In Spingomonas sp.…”