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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.


MscS, the structure and its mutations affecting cell shape.A. Crystal structure of the MscS homoheptamer (PDB ID:2OAR) representing the nonconductive state of the channel [36]. ABDOM (aa 175–265) of each subunit is shown in orange and the C-terminal fragment (aa 266–286), which is deleted in the MscSΔ266–286 mutant, is shown in red. White arrow indicates the thickness of the membrane. B. Overexpression of ABDOM leads to cell filamentation, while cells expressing wt-MscS did not differ in shape from control cells (empty vector). The highest level of cell filamentation was observed when truncated MscSΔ266–286 was expressed (note the presence of branched filaments). The amount of regular cell filaments induced by MscSΔ266–286 was unaffected by double mutation A51N/F68N, which prevents ion conduction through the channel pore. Scale bar represents 10 μm. Expression of the proteins was confirmed by Western blotting variants tagged with HA epitope (S1 Fig).
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pone.0127029.g001: MscS, the structure and its mutations affecting cell shape.A. Crystal structure of the MscS homoheptamer (PDB ID:2OAR) representing the nonconductive state of the channel [36]. ABDOM (aa 175–265) of each subunit is shown in orange and the C-terminal fragment (aa 266–286), which is deleted in the MscSΔ266–286 mutant, is shown in red. White arrow indicates the thickness of the membrane. B. Overexpression of ABDOM leads to cell filamentation, while cells expressing wt-MscS did not differ in shape from control cells (empty vector). The highest level of cell filamentation was observed when truncated MscSΔ266–286 was expressed (note the presence of branched filaments). The amount of regular cell filaments induced by MscSΔ266–286 was unaffected by double mutation A51N/F68N, which prevents ion conduction through the channel pore. Scale bar represents 10 μm. Expression of the proteins was confirmed by Western blotting variants tagged with HA epitope (S1 Fig).

Mentions: MscL has relatively simple closed—open—closed transitions during gating [5]. In contrast, after opening, the MscS channel undergoes inactivation that completely shuts the channel under sustained force [6,7]. As patch-clamp experiments show, a release of tension is a prerequisite for a return transition from the inactivated to the closed state for MscS [7,8]. Important insight into the molecular mechanism of gating has emerged from crystal structures of MscS, which have provided information on the inactivated [9] and partially open [10] conformations, as well as from kinetic analysis, electron paramagnetic resonance (EPR) studies and computational studies (reviewed in [11]). While these works led to an understanding of the conformational changes of the transmembrane domains and the force transmission between lipids and the channel, as well as solute transport through the gate, less is known about the large cytoplasmic (aa 133–286) region of MscS [9] (Fig 1, panel A). The cytoplasmic part of MscS forms a structure resembling a chamber with seven openings at the side and one at the bottom. The chamber is believed to serve as a molecular sieve [9,12,13], however, it does not form a rigid structure, as one might expect of a sieve. Instead, as demonstrated by experiments and molecular dynamics simulations, the chamber undergoes large conformational changes during channel activation, inactivation and closing, which change its shape and volume [14–20].


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)

MscS, the structure and its mutations affecting cell shape.A. Crystal structure of the MscS homoheptamer (PDB ID:2OAR) representing the nonconductive state of the channel [36]. ABDOM (aa 175–265) of each subunit is shown in orange and the C-terminal fragment (aa 266–286), which is deleted in the MscSΔ266–286 mutant, is shown in red. White arrow indicates the thickness of the membrane. B. Overexpression of ABDOM leads to cell filamentation, while cells expressing wt-MscS did not differ in shape from control cells (empty vector). The highest level of cell filamentation was observed when truncated MscSΔ266–286 was expressed (note the presence of branched filaments). The amount of regular cell filaments induced by MscSΔ266–286 was unaffected by double mutation A51N/F68N, which prevents ion conduction through the channel pore. Scale bar represents 10 μm. Expression of the proteins was confirmed by Western blotting variants tagged with HA epitope (S1 Fig).
© Copyright Policy
Related In: Results  -  Collection

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pone.0127029.g001: MscS, the structure and its mutations affecting cell shape.A. Crystal structure of the MscS homoheptamer (PDB ID:2OAR) representing the nonconductive state of the channel [36]. ABDOM (aa 175–265) of each subunit is shown in orange and the C-terminal fragment (aa 266–286), which is deleted in the MscSΔ266–286 mutant, is shown in red. White arrow indicates the thickness of the membrane. B. Overexpression of ABDOM leads to cell filamentation, while cells expressing wt-MscS did not differ in shape from control cells (empty vector). The highest level of cell filamentation was observed when truncated MscSΔ266–286 was expressed (note the presence of branched filaments). The amount of regular cell filaments induced by MscSΔ266–286 was unaffected by double mutation A51N/F68N, which prevents ion conduction through the channel pore. Scale bar represents 10 μm. Expression of the proteins was confirmed by Western blotting variants tagged with HA epitope (S1 Fig).
Mentions: MscL has relatively simple closed—open—closed transitions during gating [5]. In contrast, after opening, the MscS channel undergoes inactivation that completely shuts the channel under sustained force [6,7]. As patch-clamp experiments show, a release of tension is a prerequisite for a return transition from the inactivated to the closed state for MscS [7,8]. Important insight into the molecular mechanism of gating has emerged from crystal structures of MscS, which have provided information on the inactivated [9] and partially open [10] conformations, as well as from kinetic analysis, electron paramagnetic resonance (EPR) studies and computational studies (reviewed in [11]). While these works led to an understanding of the conformational changes of the transmembrane domains and the force transmission between lipids and the channel, as well as solute transport through the gate, less is known about the large cytoplasmic (aa 133–286) region of MscS [9] (Fig 1, panel A). The cytoplasmic part of MscS forms a structure resembling a chamber with seven openings at the side and one at the bottom. The chamber is believed to serve as a molecular sieve [9,12,13], however, it does not form a rigid structure, as one might expect of a sieve. Instead, as demonstrated by experiments and molecular dynamics simulations, the chamber undergoes large conformational changes during channel activation, inactivation and closing, which change its shape and volume [14–20].

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.