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Intracellular vesicles as reproduction elements in cell wall-deficient L-form bacteria.

Briers Y, Staubli T, Schmid MC, Wagner M, Schuppler M, Loessner MJ - PLoS ONE (2012)

Bottom Line: Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release.Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition.Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species.

View Article: PubMed Central - PubMed

Affiliation: Institute of Food, Nutrition and Health, ETH Zurich, Zurich, Switzerland.

ABSTRACT
Cell wall-deficient bacteria, or L-forms, represent an extreme example of bacterial plasticity. Stable L-forms can multiply and propagate indefinitely in the absence of a cell wall. Data presented here are consistent with the model that intracellular vesicles in Listeria monocytogenes L-form cells represent the actual viable reproductive elements. First, small intracellular vesicles are formed along the mother cell cytoplasmic membrane, originating from local phospholipid accumulation. During growth, daughter vesicles incorporate a small volume of the cellular cytoplasm, and accumulate within volume-expanding mother cells. Confocal Raman microspectroscopy demonstrated the presence of nucleic acids and proteins in all intracellular vesicles, but only a fraction of which reveals metabolic activity. Following collapse of the mother cell and release of the daughter vesicles, they can establish their own membrane potential required for respiratory and metabolic processes. Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release. Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition. Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species. From a more general viewpoint, this type of multiplication mechanism seems reminiscent of the physicochemical self-reproducing properties of abiotic lipid vesicles used to study the primordial reproduction pathways of putative prokaryotic precursor cells.

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Origin of intracellular vesicles.(A) First, uniformity of the unilamellar membrane structure is disturbed and phospholipids accumulate along the membrane. (B) Small intracellular vesicles are always generated in direct proximity to these lipid domains (left), and cytoplasmic material of the maternal cell is encapsulated into the vesicles as visualized by the compartmentalization of GFP (right). Membranes stained with CellTrace BODIPY TR methyl ester and cytoplasmic GFP fluorescence are shown in red and green, respectively. All images are confocal. (D) Current model for vesicle genesis. Lipid accumulations act as a pool for incorporation of phospholipids into a membrane structure, resulting in enlargement of intracellular progeny vesicles by a self-organizing, spontaneous process typical for bipolar phospholipids.
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pone-0038514-g002: Origin of intracellular vesicles.(A) First, uniformity of the unilamellar membrane structure is disturbed and phospholipids accumulate along the membrane. (B) Small intracellular vesicles are always generated in direct proximity to these lipid domains (left), and cytoplasmic material of the maternal cell is encapsulated into the vesicles as visualized by the compartmentalization of GFP (right). Membranes stained with CellTrace BODIPY TR methyl ester and cytoplasmic GFP fluorescence are shown in red and green, respectively. All images are confocal. (D) Current model for vesicle genesis. Lipid accumulations act as a pool for incorporation of phospholipids into a membrane structure, resulting in enlargement of intracellular progeny vesicles by a self-organizing, spontaneous process typical for bipolar phospholipids.

Mentions: To provide more insight into the possible origin and development of intracellular vesicles, we focused on the smallest size vesicles. With the aid of a membrane stain, many sites of disrupted uni-lamellarity and phospholipids accumulation could be observed along the mother cell membranes (Figure 2A). Small intracellular vesicles were always located in close proximity to these lipid accumulations (Figure 2B). Their inside consisted of an apparently normal mother cell cytoplasm, as visualized by the compartmentalization of cytosolic GFP (Figure 2C) and other proteins and nucleic acids (see below). We propose that these lipid accumulations represent the phospholipid pool and nucleation site for the generation of new vesicles. This process appears to be largely self-organizing and spontaneous, similar to the formation of liposomes. Subsequent enlargement of the vesicles may be supported by the existing lipids from the focal accumulations, or newly synthesized phospholipids from the mother cell (Figure 2D). Based on our observations, we also suggest that the development of new, small intracellular vesicles occurs in a random fashion, as the biogenesis of different intracellular vesicles often takes place simultaneously at apparently random sites along the mother cell membrane.


Intracellular vesicles as reproduction elements in cell wall-deficient L-form bacteria.

Briers Y, Staubli T, Schmid MC, Wagner M, Schuppler M, Loessner MJ - PLoS ONE (2012)

Origin of intracellular vesicles.(A) First, uniformity of the unilamellar membrane structure is disturbed and phospholipids accumulate along the membrane. (B) Small intracellular vesicles are always generated in direct proximity to these lipid domains (left), and cytoplasmic material of the maternal cell is encapsulated into the vesicles as visualized by the compartmentalization of GFP (right). Membranes stained with CellTrace BODIPY TR methyl ester and cytoplasmic GFP fluorescence are shown in red and green, respectively. All images are confocal. (D) Current model for vesicle genesis. Lipid accumulations act as a pool for incorporation of phospholipids into a membrane structure, resulting in enlargement of intracellular progeny vesicles by a self-organizing, spontaneous process typical for bipolar phospholipids.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3368840&req=5

pone-0038514-g002: Origin of intracellular vesicles.(A) First, uniformity of the unilamellar membrane structure is disturbed and phospholipids accumulate along the membrane. (B) Small intracellular vesicles are always generated in direct proximity to these lipid domains (left), and cytoplasmic material of the maternal cell is encapsulated into the vesicles as visualized by the compartmentalization of GFP (right). Membranes stained with CellTrace BODIPY TR methyl ester and cytoplasmic GFP fluorescence are shown in red and green, respectively. All images are confocal. (D) Current model for vesicle genesis. Lipid accumulations act as a pool for incorporation of phospholipids into a membrane structure, resulting in enlargement of intracellular progeny vesicles by a self-organizing, spontaneous process typical for bipolar phospholipids.
Mentions: To provide more insight into the possible origin and development of intracellular vesicles, we focused on the smallest size vesicles. With the aid of a membrane stain, many sites of disrupted uni-lamellarity and phospholipids accumulation could be observed along the mother cell membranes (Figure 2A). Small intracellular vesicles were always located in close proximity to these lipid accumulations (Figure 2B). Their inside consisted of an apparently normal mother cell cytoplasm, as visualized by the compartmentalization of cytosolic GFP (Figure 2C) and other proteins and nucleic acids (see below). We propose that these lipid accumulations represent the phospholipid pool and nucleation site for the generation of new vesicles. This process appears to be largely self-organizing and spontaneous, similar to the formation of liposomes. Subsequent enlargement of the vesicles may be supported by the existing lipids from the focal accumulations, or newly synthesized phospholipids from the mother cell (Figure 2D). Based on our observations, we also suggest that the development of new, small intracellular vesicles occurs in a random fashion, as the biogenesis of different intracellular vesicles often takes place simultaneously at apparently random sites along the mother cell membrane.

Bottom Line: Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release.Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition.Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species.

View Article: PubMed Central - PubMed

Affiliation: Institute of Food, Nutrition and Health, ETH Zurich, Zurich, Switzerland.

ABSTRACT
Cell wall-deficient bacteria, or L-forms, represent an extreme example of bacterial plasticity. Stable L-forms can multiply and propagate indefinitely in the absence of a cell wall. Data presented here are consistent with the model that intracellular vesicles in Listeria monocytogenes L-form cells represent the actual viable reproductive elements. First, small intracellular vesicles are formed along the mother cell cytoplasmic membrane, originating from local phospholipid accumulation. During growth, daughter vesicles incorporate a small volume of the cellular cytoplasm, and accumulate within volume-expanding mother cells. Confocal Raman microspectroscopy demonstrated the presence of nucleic acids and proteins in all intracellular vesicles, but only a fraction of which reveals metabolic activity. Following collapse of the mother cell and release of the daughter vesicles, they can establish their own membrane potential required for respiratory and metabolic processes. Premature depolarization of the surrounding membrane promotes activation of daughter cell metabolism prior to release. Based on genome resequencing of L-forms and comparison to the parental strain, we found no evidence for predisposing mutations that might be required for L-form transition. Further investigations revealed that propagation by intracellular budding not only occurs in Listeria species, but also in L-form cells generated from different Enterococcus species. From a more general viewpoint, this type of multiplication mechanism seems reminiscent of the physicochemical self-reproducing properties of abiotic lipid vesicles used to study the primordial reproduction pathways of putative prokaryotic precursor cells.

Show MeSH
Related in: MedlinePlus