Limits...
The essential peptidoglycan glycosyltransferase MurG forms a complex with proteins involved in lateral envelope growth as well as with proteins involved in cell division in Escherichia coli.

Mohammadi T, Karczmarek A, Crouvoisier M, Bouhss A, Mengin-Lecreulx D, den Blaauwen T - Mol. Microbiol. (2007)

Bottom Line: In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization.Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation.A model representing the first complex is proposed.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, Kruislaan 316, 1098 SM Amsterdam, PO Box 194062, 1090 GB Amsterdam, The Netherlands.

ABSTRACT
In Escherichia coli many enzymes including MurG are directly involved in the synthesis and assembly of peptidoglycan. MurG is an essential glycosyltransferase catalysing the last intracellular step of peptidoglycan synthesis. To elucidate its role during elongation and division events, localization of MurG using immunofluorescence microscopy was performed. MurG exhibited a random distribution in the cell envelope with a relatively higher intensity at the division site. This mid-cell localization was dependent on the presence of a mature divisome. Its localization in the lateral cell wall appeared to require the presence of MreCD. This could be indicative of a potential interaction between MurG and other proteins. Investigating this by immunoprecipitation revealed the association of MurG with MreB and MraY in the same protein complex. In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization. Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation. It is postulated that the involvement of MurG in the peptidoglycan synthesis concurs with two complexes, one implicated in cell elongation and the other in division. A model representing the first complex is proposed.

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MurG and MreB localization are independent of the spherical cell morphology (A and B). LMC500 cells were grown for 2 MDs in the presence of mecillinam (inhibitor of PBP2), at 28°C in GB1. Cells were immunolabelled with anti-MreB (A) or anti-MurG (B). The MreB helical structure is preserved and the multi-foci localization pattern of MurG is observed. Arrangement of MurG is dependent on the presence of MreCD (C). IFM was performed with PA340-678pMEW1 strain (mreBCD deletion strain with the pMEW1 plasmid that expresses MreC and MreD constitutively) grown at 28°C in TY to mid-exponential phase. In these spherical cells MurG localized normally as multiple foci in the cell envelope and at mid-cell. Phase contrast (left) and fluorescence images (right) are shown. Scale bar equals 1 μm.
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fig07: MurG and MreB localization are independent of the spherical cell morphology (A and B). LMC500 cells were grown for 2 MDs in the presence of mecillinam (inhibitor of PBP2), at 28°C in GB1. Cells were immunolabelled with anti-MreB (A) or anti-MurG (B). The MreB helical structure is preserved and the multi-foci localization pattern of MurG is observed. Arrangement of MurG is dependent on the presence of MreCD (C). IFM was performed with PA340-678pMEW1 strain (mreBCD deletion strain with the pMEW1 plasmid that expresses MreC and MreD constitutively) grown at 28°C in TY to mid-exponential phase. In these spherical cells MurG localized normally as multiple foci in the cell envelope and at mid-cell. Phase contrast (left) and fluorescence images (right) are shown. Scale bar equals 1 μm.

Mentions: In the parental wild-type strain PA340 MurG exhibited a normal localization as multiple foci in the cell envelope (Fig. 6B, a). As PA340-678 cells are round, it is possible that the observed abnormality in MurG localization pattern was due to the aberrant cell morphology rather than to the absence of the MreB helix. To test whether the localization of MurG would be maintained in spherical cells exhibiting a helical arrangement pattern of MreB, LMC500 cells were grown for 2 MDs in the presence of 2 μg ml−1 of the PBP2 inhibitor mecillinam (Park and Burman et al., 1973). A functional PBP2 is essential for length growth and maintenance of the diameter of the cells (Den Blaauwen et al., 2003). Therefore inhibition of PBP2 by its specific inhibitor mecillinam results in spherical growth. The round cells were subsequently labelled with anti-MurG and anti-MreB. Both patterns of MreB and MurG localization were similar to those observed in rod-shape wild-type E. coli (Fig. 7A and B respectively, see also Fig. S7). This indicates that MurG does not require a rod-shape cell morphology for a proper localization, and that cytosolic localization visualized in round ΔmreBCD cells is most likely due to the absence of the Mre proteins rather than to the altered cell morphology.


The essential peptidoglycan glycosyltransferase MurG forms a complex with proteins involved in lateral envelope growth as well as with proteins involved in cell division in Escherichia coli.

Mohammadi T, Karczmarek A, Crouvoisier M, Bouhss A, Mengin-Lecreulx D, den Blaauwen T - Mol. Microbiol. (2007)

MurG and MreB localization are independent of the spherical cell morphology (A and B). LMC500 cells were grown for 2 MDs in the presence of mecillinam (inhibitor of PBP2), at 28°C in GB1. Cells were immunolabelled with anti-MreB (A) or anti-MurG (B). The MreB helical structure is preserved and the multi-foci localization pattern of MurG is observed. Arrangement of MurG is dependent on the presence of MreCD (C). IFM was performed with PA340-678pMEW1 strain (mreBCD deletion strain with the pMEW1 plasmid that expresses MreC and MreD constitutively) grown at 28°C in TY to mid-exponential phase. In these spherical cells MurG localized normally as multiple foci in the cell envelope and at mid-cell. Phase contrast (left) and fluorescence images (right) are shown. Scale bar equals 1 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2170320&req=5

fig07: MurG and MreB localization are independent of the spherical cell morphology (A and B). LMC500 cells were grown for 2 MDs in the presence of mecillinam (inhibitor of PBP2), at 28°C in GB1. Cells were immunolabelled with anti-MreB (A) or anti-MurG (B). The MreB helical structure is preserved and the multi-foci localization pattern of MurG is observed. Arrangement of MurG is dependent on the presence of MreCD (C). IFM was performed with PA340-678pMEW1 strain (mreBCD deletion strain with the pMEW1 plasmid that expresses MreC and MreD constitutively) grown at 28°C in TY to mid-exponential phase. In these spherical cells MurG localized normally as multiple foci in the cell envelope and at mid-cell. Phase contrast (left) and fluorescence images (right) are shown. Scale bar equals 1 μm.
Mentions: In the parental wild-type strain PA340 MurG exhibited a normal localization as multiple foci in the cell envelope (Fig. 6B, a). As PA340-678 cells are round, it is possible that the observed abnormality in MurG localization pattern was due to the aberrant cell morphology rather than to the absence of the MreB helix. To test whether the localization of MurG would be maintained in spherical cells exhibiting a helical arrangement pattern of MreB, LMC500 cells were grown for 2 MDs in the presence of 2 μg ml−1 of the PBP2 inhibitor mecillinam (Park and Burman et al., 1973). A functional PBP2 is essential for length growth and maintenance of the diameter of the cells (Den Blaauwen et al., 2003). Therefore inhibition of PBP2 by its specific inhibitor mecillinam results in spherical growth. The round cells were subsequently labelled with anti-MurG and anti-MreB. Both patterns of MreB and MurG localization were similar to those observed in rod-shape wild-type E. coli (Fig. 7A and B respectively, see also Fig. S7). This indicates that MurG does not require a rod-shape cell morphology for a proper localization, and that cytosolic localization visualized in round ΔmreBCD cells is most likely due to the absence of the Mre proteins rather than to the altered cell morphology.

Bottom Line: In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization.Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation.A model representing the first complex is proposed.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, Kruislaan 316, 1098 SM Amsterdam, PO Box 194062, 1090 GB Amsterdam, The Netherlands.

ABSTRACT
In Escherichia coli many enzymes including MurG are directly involved in the synthesis and assembly of peptidoglycan. MurG is an essential glycosyltransferase catalysing the last intracellular step of peptidoglycan synthesis. To elucidate its role during elongation and division events, localization of MurG using immunofluorescence microscopy was performed. MurG exhibited a random distribution in the cell envelope with a relatively higher intensity at the division site. This mid-cell localization was dependent on the presence of a mature divisome. Its localization in the lateral cell wall appeared to require the presence of MreCD. This could be indicative of a potential interaction between MurG and other proteins. Investigating this by immunoprecipitation revealed the association of MurG with MreB and MraY in the same protein complex. In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization. Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation. It is postulated that the involvement of MurG in the peptidoglycan synthesis concurs with two complexes, one implicated in cell elongation and the other in division. A model representing the first complex is proposed.

Show MeSH
Related in: MedlinePlus