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Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

Koprowski P, Grajkowski W, Balcerzak M, Filipiuk I, Fabczak H, Kubalski A - PLoS ONE (2015)

Bottom Line: MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel.Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein.Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Pasteur 3, Warsaw, Poland.

ABSTRACT
Bacterial mechano-sensitive (MS) channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS) family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

No MeSH data available.


Related in: MedlinePlus

Hypothetical model of MscS acting as a dual-action protective gate.A. Under non-stress conditions MscS remains in the closed state and the ABDOM domain of MscS is inaccessible to interaction with FtsZ. B. Under severe osmotic downshocks the channel opens and jettisons osmolytes rapidly returning to the closed state. The short-lived open state does not result in formation of the cell-wall remodeling complex. C. Under slowly rising and prolonged local increase of tension within the bulges of the membrane the channel performs transition into the inactivated state. In this channel conformation ABDOM becomes accessible and binds FtsZ.
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pone.0127029.g005: Hypothetical model of MscS acting as a dual-action protective gate.A. Under non-stress conditions MscS remains in the closed state and the ABDOM domain of MscS is inaccessible to interaction with FtsZ. B. Under severe osmotic downshocks the channel opens and jettisons osmolytes rapidly returning to the closed state. The short-lived open state does not result in formation of the cell-wall remodeling complex. C. Under slowly rising and prolonged local increase of tension within the bulges of the membrane the channel performs transition into the inactivated state. In this channel conformation ABDOM becomes accessible and binds FtsZ.

Mentions: We have demonstrated that MscS protects cells against antibiotic stress and this function is independent of its ability to protect against osmotic downshocks. We believe that MscS acts here as an interpreter of local changes of membrane tension due to small lesions induced by β-lactam antibiotics. The question arises as to how MscS channel can sense local envelope stress. It is well established that Gram-negative PG is a single layer mesh formed by long strands of glycans cross-linked by stretchable peptides [53–56]. This forms a structure with pores known as “tesserae” [55]. The existing models predict that breaking peptide cross-links leads to defects which eventually end up with cell lysis. However, before it occurs the turgor pressure expands the envelope around the defects to form “bulges”. Such bulges or blebs have been found in in the midcell of cells treated with β-lactam antibiotics [37,57,58] but also cells undergoing slow osmotic downshocks [59] indicating for susceptibility of the cell wall at the division site. It is very likely that the membrane of the bulges contains MscS protein. In fact it was demonstrated that MscS prefers to localize in an osmotic-stress—dependent manner at cell poles in cardiolipin (CL) [60]. CL shows preference to curved membranes and it was shown that CL forms micro-domains in E. coli membranes in response to curvature [61]. It is therefore reasonable to assume that bulges will contain CL micro-domains which in turn could result in preferential localization of MscS in the bulges. It is well established that MscS gates in response to tension and curvature [11], which are different in the bulge than in the rest of the cell, and the specific localization of MscS allows the channel to sense tension only locally, and possibly, to direct FtsZ for local PG repair. Other data also indicate that tension affects PG synthesis: i. the curved rod shape of Caulobacter crescentus depends on the presence of intracellular filament of crescentin attached to the membrane [62], ii. crescentin expressed in E. coli makes the cells curved as a result of crescentin-dependent gradient of PG insertion around the cell circumference [63]. In our model we propose that MscS plays active role by recruiting FtsZ to the sites of damage, what in turn can direct cell wall synthesi/repair (Fig 5). However, we cannot exclude entirely the possibility that the role of MscS is passive and local binding of FtsZ blocks division as long as the cell wall is not repaired by an independent mechanism. This could be supported by observations that blocking of cell division has protective effects when E. coli cells are treated with some β-lactams [39,64,65]. Despite the fact that we do not know exact mechanism by which MscS cooperates with FtsZ to fulfill cellular function, we believe that MscS-FtsZ interaction is important in specific stress conditions. We have to note that we were not able to prove that native expression of MscS protects against β-lactams (not shown). However, the effect could be subtle and masked by other mechanisms. Similarly, there are multiple MscS paralogs of MscS in E. coli and initial experiments suggested that they cannot protect cells against osmotic downshocks unless overexpressed [66]. However, recently it was found that the protection provided by MscS homologs depends strongly on the rate of osmotic downshock and under a slow enough osmotic drop, MscS homologs can lead to survival rates comparable to those found in wild-type strains [67].


Cytoplasmic Domain of MscS Interacts with Cell Division Protein FtsZ: A Possible Non-Channel Function of the Mechanosensitive Channel in Escherichia Coli.

Koprowski P, Grajkowski W, Balcerzak M, Filipiuk I, Fabczak H, Kubalski A - PLoS ONE (2015)

Hypothetical model of MscS acting as a dual-action protective gate.A. Under non-stress conditions MscS remains in the closed state and the ABDOM domain of MscS is inaccessible to interaction with FtsZ. B. Under severe osmotic downshocks the channel opens and jettisons osmolytes rapidly returning to the closed state. The short-lived open state does not result in formation of the cell-wall remodeling complex. C. Under slowly rising and prolonged local increase of tension within the bulges of the membrane the channel performs transition into the inactivated state. In this channel conformation ABDOM becomes accessible and binds FtsZ.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4440785&req=5

pone.0127029.g005: Hypothetical model of MscS acting as a dual-action protective gate.A. Under non-stress conditions MscS remains in the closed state and the ABDOM domain of MscS is inaccessible to interaction with FtsZ. B. Under severe osmotic downshocks the channel opens and jettisons osmolytes rapidly returning to the closed state. The short-lived open state does not result in formation of the cell-wall remodeling complex. C. Under slowly rising and prolonged local increase of tension within the bulges of the membrane the channel performs transition into the inactivated state. In this channel conformation ABDOM becomes accessible and binds FtsZ.
Mentions: We have demonstrated that MscS protects cells against antibiotic stress and this function is independent of its ability to protect against osmotic downshocks. We believe that MscS acts here as an interpreter of local changes of membrane tension due to small lesions induced by β-lactam antibiotics. The question arises as to how MscS channel can sense local envelope stress. It is well established that Gram-negative PG is a single layer mesh formed by long strands of glycans cross-linked by stretchable peptides [53–56]. This forms a structure with pores known as “tesserae” [55]. The existing models predict that breaking peptide cross-links leads to defects which eventually end up with cell lysis. However, before it occurs the turgor pressure expands the envelope around the defects to form “bulges”. Such bulges or blebs have been found in in the midcell of cells treated with β-lactam antibiotics [37,57,58] but also cells undergoing slow osmotic downshocks [59] indicating for susceptibility of the cell wall at the division site. It is very likely that the membrane of the bulges contains MscS protein. In fact it was demonstrated that MscS prefers to localize in an osmotic-stress—dependent manner at cell poles in cardiolipin (CL) [60]. CL shows preference to curved membranes and it was shown that CL forms micro-domains in E. coli membranes in response to curvature [61]. It is therefore reasonable to assume that bulges will contain CL micro-domains which in turn could result in preferential localization of MscS in the bulges. It is well established that MscS gates in response to tension and curvature [11], which are different in the bulge than in the rest of the cell, and the specific localization of MscS allows the channel to sense tension only locally, and possibly, to direct FtsZ for local PG repair. Other data also indicate that tension affects PG synthesis: i. the curved rod shape of Caulobacter crescentus depends on the presence of intracellular filament of crescentin attached to the membrane [62], ii. crescentin expressed in E. coli makes the cells curved as a result of crescentin-dependent gradient of PG insertion around the cell circumference [63]. In our model we propose that MscS plays active role by recruiting FtsZ to the sites of damage, what in turn can direct cell wall synthesi/repair (Fig 5). However, we cannot exclude entirely the possibility that the role of MscS is passive and local binding of FtsZ blocks division as long as the cell wall is not repaired by an independent mechanism. This could be supported by observations that blocking of cell division has protective effects when E. coli cells are treated with some β-lactams [39,64,65]. Despite the fact that we do not know exact mechanism by which MscS cooperates with FtsZ to fulfill cellular function, we believe that MscS-FtsZ interaction is important in specific stress conditions. We have to note that we were not able to prove that native expression of MscS protects against β-lactams (not shown). However, the effect could be subtle and masked by other mechanisms. Similarly, there are multiple MscS paralogs of MscS in E. coli and initial experiments suggested that they cannot protect cells against osmotic downshocks unless overexpressed [66]. However, recently it was found that the protection provided by MscS homologs depends strongly on the rate of osmotic downshock and under a slow enough osmotic drop, MscS homologs can lead to survival rates comparable to those found in wild-type strains [67].

Bottom Line: MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel.Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein.Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology, Nencki Institute of Experimental Biology, Polish Academy of Sciences, Pasteur 3, Warsaw, Poland.

ABSTRACT
Bacterial mechano-sensitive (MS) channels reside in the inner membrane and are considered to act as emergency valves whose role is to lower cell turgor when bacteria enter hypo-osmotic environments. However, there is emerging evidence that members of the Mechano-sensitive channel Small (MscS) family play additional roles in bacterial and plant cell physiology. MscS has a large cytoplasmic C-terminal region that changes its shape upon activation and inactivation of the channel. Our pull-down and co-sedimentation assays show that this domain interacts with FtsZ, a bacterial tubulin-like protein. We identify point mutations in the MscS C-terminal domain that reduce binding to FtsZ and show that bacteria expressing these mutants are compromised in growth on sublethal concentrations of β-lactam antibiotics. Our results suggest that interaction between MscS and FtsZ could occur upon inactivation and/or opening of the channel and could be important for the bacterial cell response against sustained stress upon stationary phase and in the presence of β-lactam antibiotics.

No MeSH data available.


Related in: MedlinePlus