Synthesis, Release, and Recapture of Compatible Solute Proline by Osmotically Stressed Bacillus subtilis Cells

Abstract

Bacillus subtilis synthesizes large amounts of the compatible solute proline as a cellular defense against high osmolarity to ensure a physiologically appropriate level of hydration of the cytoplasm and turgor. It also imports proline for this purpose via the osmotically inducible OpuE transport system. Unexpectedly, an opuE mutant was at a strong growth disadvantage in high-salinity minimal media lacking proline. Appreciable amounts of proline were detected in the culture supernatant of the opuE mutant strain, and they rose concomitantly with increases in the external salinity. We found that the intracellular proline pool of severely salinity-stressed cells of the opuE mutant was considerably lower than that of its opuE ؉ parent strain. This loss of proline into the medium and the resulting decrease in the intracellular proline content provide a rational explanation for the observed salt-sensitive growth phenotype of cells lacking OpuE. None of the known MscL-and MscS-type mechanosensitive channels of B. subtilis participated in the release of proline under permanently imposed high-salinity growth conditions. The data reported here show that the OpuE transporter not only possesses the previously reported role for the scavenging of exogenously provided proline as an osmoprotectant but also functions as a physiologically highly important recapturing device for proline that is synthesized de novo and subsequently released by salt-stressed B. subtilis cells. The wider implications of our findings for the retention of compatible solutes by osmotically challenged microorganisms and the roles of uptake systems for compatible solutes are considered.

Osmotic Control of opuA Expression in Bacillus subtilis and Its Modulation in Response to Intracellular Glycine Betaine and Proline Pools

“…Release of glycine betaine by a MscS-type channel has been observed in Corynebacterium glutamicum (87), and such channels also exist in B. subtilis (88,89). However, any role of MscL-or MscS-type channels in the release of proline from continuously osmotically challenged B. subtilis cells has been ruled out (84).…”

“…A process of release and recapture of compatible solutes by continuously osmotically stressed B. subtilis cells, which we have recently discovered for newly produced proline (84), also exists for glycine betaine synthesized from the precursor choline (S. Moses and E. Bremer, unpublished data). This activity will aid the cell in finely tuning the glycine betaine pools at a given salinity.…”

Homeostasis and Catabolism of Choline and Glycine Betaine: Lessons from Pseudomonas aeruginosa

“…In a mechanistic examination of this phenomenon, Hoffmann and colleagues recently reported the sharing and recycling of proline between cells of Bacillus subtilis, providing direct evidence of osmoprotectants as a piece of microbial communal property (103). In the same report, Hoffmann et al communicate that B. subtilis is capable of sharing GB within a population (103). It remains to be demonstrated what impact such resource sharing of GB has on intraspecies survival or the extent to which interspecies sharing of GB occurs within microbial communities beyond Vibrio.…”

The γ-Aminobutyrate Permease GabP Serves as the Third Proline Transporter of Bacillus subtilis

“…Their primary physiological roles can be discerned from the distinct transcriptional profile of their structural genes (10,16,23,28,29) and the different abilities of the PutP and OpuE proteins to withstand the inhibiting effects of high salinity on their transport activity (10). These features associate PutP with the proline catabolic system of B. subtilis (10,28,29) and OpuE with the cell's osmostress response systems (16,22,72,73). Nevertheless, it is worth noting that our data now show not only that the OpuE transporter imports proline when it is used as an osmoprotectant (15,16) but also that it can do so when B. subtilis uses it as a nutrient (see Fig.…”

“…Proline-mediated osmoprotection can also be achieved through uptake; the osmotically inducible OpuE transporter of B. subtilis serves this function (15,16). OpuE also operates as a recapturing device for newly synthesized proline that is released from B. subtilis grown under high-salinity conditions, probably in an effort by the cell to fine-tune turgor (22). Expression of opuE is osmotically inducible (16,23,24), but there is no stimulation of opuE expression in response to proline availability.…”