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Contact- and Protein Transfer-Dependent Stimulation of Assembly of the Gliding Motility Machinery in Myxococcus xanthus.

Jakobczak B, Keilberg D, Wuichet K, Søgaard-Andersen L - PLoS Genet. (2015)

Bottom Line: Conversely, incorporation of AglZ and AglQ into the gliding motility complex depends on CglC, GltB, GltA and GltC.Remarkably, physical transfer of the OM lipoprotein CglC to a ΔcglC recipient stimulates assembly of the gliding motility complex in the recipient likely by facilitating the OM integration of GltB and GltA.These data provide evidence that the gliding motility complex in M. xanthus includes OM proteins and suggest that this complex extends from the cytoplasm across the cell envelope to the OM.

View Article: PubMed Central - PubMed

Affiliation: Department of Ecophysiology, Max Planck Institute for Terrestrial Microbiology, Marburg, Germany.

ABSTRACT
Bacteria engage in contact-dependent activities to coordinate cellular activities that aid their survival. Cells of Myxococcus xanthus move over surfaces by means of type IV pili and gliding motility. Upon direct contact, cells physically exchange outer membrane (OM) lipoproteins, and this transfer can rescue motility in mutants lacking lipoproteins required for motility. The mechanism of gliding motility and its stimulation by transferred OM lipoproteins remain poorly characterized. We investigated the function of CglC, GltB, GltA and GltC, all of which are required for gliding. We demonstrate that CglC is an OM lipoprotein, GltB and GltA are integral OM β-barrel proteins, and GltC is a soluble periplasmic protein. GltB and GltA are mutually stabilizing, and both are required to stabilize GltC, whereas CglC accumulate independently of GltB, GltA and GltC. Consistently, purified GltB, GltA and GltC proteins interact in all pair-wise combinations. Using active fluorescently-tagged fusion proteins, we demonstrate that GltB, GltA and GltC are integral components of the gliding motility complex. Incorporation of GltB and GltA into this complex depends on CglC and GltC as well as on the cytoplasmic AglZ protein and the inner membrane protein AglQ, both of which are components of the gliding motility complex. Conversely, incorporation of AglZ and AglQ into the gliding motility complex depends on CglC, GltB, GltA and GltC. Remarkably, physical transfer of the OM lipoprotein CglC to a ΔcglC recipient stimulates assembly of the gliding motility complex in the recipient likely by facilitating the OM integration of GltB and GltA. These data provide evidence that the gliding motility complex in M. xanthus includes OM proteins and suggest that this complex extends from the cytoplasm across the cell envelope to the OM. These data add assembly of gliding motility complexes in M. xanthus to the growing list of contact-dependent activities in bacteria.

No MeSH data available.


Related in: MedlinePlus

Subcellular localization of proteins required for gliding motility.Synonyms for proteins required for gliding motility are indicated on the left. Proteins shown on a grey, brown or purple background in the left panel and in grey, brown or purple in the right panel are encoded together in the genome [17]; proteins in white are not encoded near other proteins shown here. Proteins outlined in red have been shown by fluorescence microscopy to localize in clusters along the cell body [here; [15–18]]; proteins that interact based on pull down experiments using M. xanthus cell extracts are indicated in italics [15]. CglB is an OM lipoprotein [29] and CglD is predicted to be an OM lipoprotein [20]. It is not known if they face towards the periplasm or are exposed on the cell surface. They are shown on the cell surface because they contain a von Willebrand domain (VWA_2), which is often involved in cell adhesion [30].
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pgen.1005341.g001: Subcellular localization of proteins required for gliding motility.Synonyms for proteins required for gliding motility are indicated on the left. Proteins shown on a grey, brown or purple background in the left panel and in grey, brown or purple in the right panel are encoded together in the genome [17]; proteins in white are not encoded near other proteins shown here. Proteins outlined in red have been shown by fluorescence microscopy to localize in clusters along the cell body [here; [15–18]]; proteins that interact based on pull down experiments using M. xanthus cell extracts are indicated in italics [15]. CglB is an OM lipoprotein [29] and CglD is predicted to be an OM lipoprotein [20]. It is not known if they face towards the periplasm or are exposed on the cell surface. They are shown on the cell surface because they contain a von Willebrand domain (VWA_2), which is often involved in cell adhesion [30].

Mentions: Numerous proteins involved in gliding motility have been described [13,17,18,20–22]. Genetic and cytological evidence suggests that gliding motility is driven by a protein complex that spans part or all of the cell envelope. This complex includes the AglR, AglQ and AglS proteins, which are homologs of MotA/TolQ/ExbB (AglR) and MotB/TolR/ExbD (AglQ and AglS) and form a proton channel in the inner membrane (IM) [13,18]. AglQ and AglR have been shown directly to localize to the clusters of motility complexes [13,23] (Fig 1). Additional proteins that localize to the cytoplasm, IM, periplasm or outer membrane (OM) have been suggested to be components of the gliding motility complex. These proteins include the 11 GltA-K proteins that are encoded by two gene clusters (Fig 1) and among which the eight GltA-H are paralogs of the NfsA-H proteins that are important for spore formation [17,24–27]. The NfsA-H proteins have been suggested to form a complex that spans the cell envelope and with protein localizing to the IM, periplasm and OM. Similarly, the GltA-K proteins have been suggested to form a complex that would span from the cytoplasm to the OM [17]. Several of the proteins required for gliding motility have been shown to interact based on pull down experiments [15,17] (Fig 1). However, only the cytoplasmic AglZ [16], CglF (GltF), which is reported to be a periplasmic as well as an IM protein, and GltD, which is reported to localize to the cytoplasm and periplasm, have been shown to localize to the clusters of motility complexes along the cell length [15,17] (Fig 1). GltD and AglR have also been reported to localize to a rotating helical structure [18,23]. Accordingly, two models for the gliding machinery in M. xanthus have been proposed. In both models, motility complexes are distributed along the length of the ventral side of the cell. In the focal adhesion model, the motility complex spans from the cytoplasm over the IM, periplasm, OM to the cell surface where it connects to the substratum via the slime and generate traction force [13,17,19]. In the helical rotor model, the motility complex spans the IM and periplasm and move on a rotating helix. These complexes are hypothesized to slow down close to the substratum in that way forming clusters and causing a deformation of the cell surface. These deformations, in turn, generate drag forces between the cell and the substratum [18,23]. An analysis of the biophysical properties of cell-substrate interactions during gliding, provided evidence in favor of a focal adhesion model [28]. A prediction from the focal adhesion complex model is that the gliding motility complex includes proteins that localize to the OM.


Contact- and Protein Transfer-Dependent Stimulation of Assembly of the Gliding Motility Machinery in Myxococcus xanthus.

Jakobczak B, Keilberg D, Wuichet K, Søgaard-Andersen L - PLoS Genet. (2015)

Subcellular localization of proteins required for gliding motility.Synonyms for proteins required for gliding motility are indicated on the left. Proteins shown on a grey, brown or purple background in the left panel and in grey, brown or purple in the right panel are encoded together in the genome [17]; proteins in white are not encoded near other proteins shown here. Proteins outlined in red have been shown by fluorescence microscopy to localize in clusters along the cell body [here; [15–18]]; proteins that interact based on pull down experiments using M. xanthus cell extracts are indicated in italics [15]. CglB is an OM lipoprotein [29] and CglD is predicted to be an OM lipoprotein [20]. It is not known if they face towards the periplasm or are exposed on the cell surface. They are shown on the cell surface because they contain a von Willebrand domain (VWA_2), which is often involved in cell adhesion [30].
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4488436&req=5

pgen.1005341.g001: Subcellular localization of proteins required for gliding motility.Synonyms for proteins required for gliding motility are indicated on the left. Proteins shown on a grey, brown or purple background in the left panel and in grey, brown or purple in the right panel are encoded together in the genome [17]; proteins in white are not encoded near other proteins shown here. Proteins outlined in red have been shown by fluorescence microscopy to localize in clusters along the cell body [here; [15–18]]; proteins that interact based on pull down experiments using M. xanthus cell extracts are indicated in italics [15]. CglB is an OM lipoprotein [29] and CglD is predicted to be an OM lipoprotein [20]. It is not known if they face towards the periplasm or are exposed on the cell surface. They are shown on the cell surface because they contain a von Willebrand domain (VWA_2), which is often involved in cell adhesion [30].
Mentions: Numerous proteins involved in gliding motility have been described [13,17,18,20–22]. Genetic and cytological evidence suggests that gliding motility is driven by a protein complex that spans part or all of the cell envelope. This complex includes the AglR, AglQ and AglS proteins, which are homologs of MotA/TolQ/ExbB (AglR) and MotB/TolR/ExbD (AglQ and AglS) and form a proton channel in the inner membrane (IM) [13,18]. AglQ and AglR have been shown directly to localize to the clusters of motility complexes [13,23] (Fig 1). Additional proteins that localize to the cytoplasm, IM, periplasm or outer membrane (OM) have been suggested to be components of the gliding motility complex. These proteins include the 11 GltA-K proteins that are encoded by two gene clusters (Fig 1) and among which the eight GltA-H are paralogs of the NfsA-H proteins that are important for spore formation [17,24–27]. The NfsA-H proteins have been suggested to form a complex that spans the cell envelope and with protein localizing to the IM, periplasm and OM. Similarly, the GltA-K proteins have been suggested to form a complex that would span from the cytoplasm to the OM [17]. Several of the proteins required for gliding motility have been shown to interact based on pull down experiments [15,17] (Fig 1). However, only the cytoplasmic AglZ [16], CglF (GltF), which is reported to be a periplasmic as well as an IM protein, and GltD, which is reported to localize to the cytoplasm and periplasm, have been shown to localize to the clusters of motility complexes along the cell length [15,17] (Fig 1). GltD and AglR have also been reported to localize to a rotating helical structure [18,23]. Accordingly, two models for the gliding machinery in M. xanthus have been proposed. In both models, motility complexes are distributed along the length of the ventral side of the cell. In the focal adhesion model, the motility complex spans from the cytoplasm over the IM, periplasm, OM to the cell surface where it connects to the substratum via the slime and generate traction force [13,17,19]. In the helical rotor model, the motility complex spans the IM and periplasm and move on a rotating helix. These complexes are hypothesized to slow down close to the substratum in that way forming clusters and causing a deformation of the cell surface. These deformations, in turn, generate drag forces between the cell and the substratum [18,23]. An analysis of the biophysical properties of cell-substrate interactions during gliding, provided evidence in favor of a focal adhesion model [28]. A prediction from the focal adhesion complex model is that the gliding motility complex includes proteins that localize to the OM.

Bottom Line: Conversely, incorporation of AglZ and AglQ into the gliding motility complex depends on CglC, GltB, GltA and GltC.Remarkably, physical transfer of the OM lipoprotein CglC to a ΔcglC recipient stimulates assembly of the gliding motility complex in the recipient likely by facilitating the OM integration of GltB and GltA.These data provide evidence that the gliding motility complex in M. xanthus includes OM proteins and suggest that this complex extends from the cytoplasm across the cell envelope to the OM.

View Article: PubMed Central - PubMed

Affiliation: Department of Ecophysiology, Max Planck Institute for Terrestrial Microbiology, Marburg, Germany.

ABSTRACT
Bacteria engage in contact-dependent activities to coordinate cellular activities that aid their survival. Cells of Myxococcus xanthus move over surfaces by means of type IV pili and gliding motility. Upon direct contact, cells physically exchange outer membrane (OM) lipoproteins, and this transfer can rescue motility in mutants lacking lipoproteins required for motility. The mechanism of gliding motility and its stimulation by transferred OM lipoproteins remain poorly characterized. We investigated the function of CglC, GltB, GltA and GltC, all of which are required for gliding. We demonstrate that CglC is an OM lipoprotein, GltB and GltA are integral OM β-barrel proteins, and GltC is a soluble periplasmic protein. GltB and GltA are mutually stabilizing, and both are required to stabilize GltC, whereas CglC accumulate independently of GltB, GltA and GltC. Consistently, purified GltB, GltA and GltC proteins interact in all pair-wise combinations. Using active fluorescently-tagged fusion proteins, we demonstrate that GltB, GltA and GltC are integral components of the gliding motility complex. Incorporation of GltB and GltA into this complex depends on CglC and GltC as well as on the cytoplasmic AglZ protein and the inner membrane protein AglQ, both of which are components of the gliding motility complex. Conversely, incorporation of AglZ and AglQ into the gliding motility complex depends on CglC, GltB, GltA and GltC. Remarkably, physical transfer of the OM lipoprotein CglC to a ΔcglC recipient stimulates assembly of the gliding motility complex in the recipient likely by facilitating the OM integration of GltB and GltA. These data provide evidence that the gliding motility complex in M. xanthus includes OM proteins and suggest that this complex extends from the cytoplasm across the cell envelope to the OM. These data add assembly of gliding motility complexes in M. xanthus to the growing list of contact-dependent activities in bacteria.

No MeSH data available.


Related in: MedlinePlus