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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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Related in: MedlinePlus

Release of internal vesicles.(A) Following collapse of an L-form mother cell, intact progeny cells are released. Membranes are visible by using DIC (Differential Interference Contrast) microscopy (left); dye-labeled membranes (right) are shown immediately before (upper) and after the collapse (lower). (B) A vesiculated cell labeled with Rho123 (upper) disintegrates and releases a vesicle (lower image, arrow head). The increase in the Rho123 signal on the surface of the released vesicle suggests an increase in membrane charge.
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pone-0038514-g004: Release of internal vesicles.(A) Following collapse of an L-form mother cell, intact progeny cells are released. Membranes are visible by using DIC (Differential Interference Contrast) microscopy (left); dye-labeled membranes (right) are shown immediately before (upper) and after the collapse (lower). (B) A vesiculated cell labeled with Rho123 (upper) disintegrates and releases a vesicle (lower image, arrow head). The increase in the Rho123 signal on the surface of the released vesicle suggests an increase in membrane charge.

Mentions: Unexpectedly, most of the intracellular vesicles released into the medium by collapse of the mother cell remain intact (Figure 4A). This process is often accompanied by an increase in Rho123 fluorescence, indicating an upshift in membrane potential upon release (Figure 4B). This suggested that intracellular vesicles are “activated” upon release from the mother cell, as shown in Figure 5A. Initially, only one of the daughter vesicles released from of a collapsed mother cell shows a green fluorescent GFP signal, while three others appear shortly thereafter, within approximately 1 minute. The delay might be due to the requirement for oxidation of the immature fluorophores of pre-synthesized, non-fluorescent GFP molecules, rather than de novo gfp transcription and translation, and may also be influenced by a pH shift within the vesicles after release from the cytoplasmic compartment. Freshly released vesicles frequently feature (initially weak) signals from both CTC reduction and GFP fluorescence (Figure 5B).


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)

Release of internal vesicles.(A) Following collapse of an L-form mother cell, intact progeny cells are released. Membranes are visible by using DIC (Differential Interference Contrast) microscopy (left); dye-labeled membranes (right) are shown immediately before (upper) and after the collapse (lower). (B) A vesiculated cell labeled with Rho123 (upper) disintegrates and releases a vesicle (lower image, arrow head). The increase in the Rho123 signal on the surface of the released vesicle suggests an increase in membrane charge.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0038514-g004: Release of internal vesicles.(A) Following collapse of an L-form mother cell, intact progeny cells are released. Membranes are visible by using DIC (Differential Interference Contrast) microscopy (left); dye-labeled membranes (right) are shown immediately before (upper) and after the collapse (lower). (B) A vesiculated cell labeled with Rho123 (upper) disintegrates and releases a vesicle (lower image, arrow head). The increase in the Rho123 signal on the surface of the released vesicle suggests an increase in membrane charge.
Mentions: Unexpectedly, most of the intracellular vesicles released into the medium by collapse of the mother cell remain intact (Figure 4A). This process is often accompanied by an increase in Rho123 fluorescence, indicating an upshift in membrane potential upon release (Figure 4B). This suggested that intracellular vesicles are “activated” upon release from the mother cell, as shown in Figure 5A. Initially, only one of the daughter vesicles released from of a collapsed mother cell shows a green fluorescent GFP signal, while three others appear shortly thereafter, within approximately 1 minute. The delay might be due to the requirement for oxidation of the immature fluorophores of pre-synthesized, non-fluorescent GFP molecules, rather than de novo gfp transcription and translation, and may also be influenced by a pH shift within the vesicles after release from the cytoplasmic compartment. Freshly released vesicles frequently feature (initially weak) signals from both CTC reduction and GFP fluorescence (Figure 5B).

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