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The role of autophagy in the intracellular survival of Campylobacter concisus.

Burgos-Portugal JA, Mitchell HM, Castaño-Rodríguez N, Kaakoush NO - FEBS Open Bio (2014)

Bottom Line: Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance.C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition.Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

View Article: PubMed Central - PubMed

Affiliation: School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, NSW 2052, Australia.

ABSTRACT
Campylobacter concisus is an emerging pathogen that has been associated with gastrointestinal diseases. Given the importance of autophagy for the elimination of intracellular bacteria and the subversion of this process by pathogenic bacteria, we investigated the role of autophagy in C. concisus intracellular survival. Gentamicin protection assays were employed to assess intracellular levels of C. concisus within Caco-2 cells, following autophagy induction and inhibition. To assess the interaction between C. concisus and autophagosomes, confocal microscopy, scanning electron microscopy, and transmission electron microscopy were employed. Expression levels of 84 genes involved in the autophagy process were measured using qPCR. Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance. C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition. Confocal microscopy showed co-localization of the bacterium with autophagosomes, while transmission electron microscopy identified intracellular bacteria persisting within autophagic vesicles. Further, qPCR showed that following infection, 13 genes involved in the autophagy process were significantly regulated, and a further five showed borderline results, with an overall indication towards a dampening effect exerted by the bacterium on this process. Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

No MeSH data available.


Related in: MedlinePlus

Visualization of the internalization of C. concisus into Caco-2 cells using transmission electron microscopy. (A) C. concisus UNSWCD invading inside Caco-2 cells shown in cross section measuring 0.5 μm wide. The bacterial cell appears to be in close proximity to a mitochondrion with a fine layer surrounding them. (B) A membrane forms around internalized C. concisus UNSWCD. A fine layer surrounds the bacterium alongside mitochondria and could be indicative of early stage phagosome formation in Caco-2 cells. (C) The appearance of fine filament arrangements close to the bacterium. (D) C. concisus UNSWCD associated with an autophagosome showing a double membrane. (E) C. concisus UNSWCD shown in cross section (0.5 μm wide) in close proximity to an intermediary phagosome. The bacterium appears to be within a vacuole. (F) C. concisus UNSWCD shown in cross section (0.5 μm wide). The bacterium is shown in close proximity with a vacuole compartment forming one large phagosome. (G) High magnification image of the bacterium fusing with a vacuole compartment. (H) C. concisus UNSWCD (0.5 μm wide) observed inside an autophagolysosome containing dense granular material. (I) A Caco-2 cell showing a clear apical membrane surface with microvilli. C. concisus UNSWCD which has maintained its spiral morphology is visualised inside an autophagolysosome.
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f0035: Visualization of the internalization of C. concisus into Caco-2 cells using transmission electron microscopy. (A) C. concisus UNSWCD invading inside Caco-2 cells shown in cross section measuring 0.5 μm wide. The bacterial cell appears to be in close proximity to a mitochondrion with a fine layer surrounding them. (B) A membrane forms around internalized C. concisus UNSWCD. A fine layer surrounds the bacterium alongside mitochondria and could be indicative of early stage phagosome formation in Caco-2 cells. (C) The appearance of fine filament arrangements close to the bacterium. (D) C. concisus UNSWCD associated with an autophagosome showing a double membrane. (E) C. concisus UNSWCD shown in cross section (0.5 μm wide) in close proximity to an intermediary phagosome. The bacterium appears to be within a vacuole. (F) C. concisus UNSWCD shown in cross section (0.5 μm wide). The bacterium is shown in close proximity with a vacuole compartment forming one large phagosome. (G) High magnification image of the bacterium fusing with a vacuole compartment. (H) C. concisus UNSWCD (0.5 μm wide) observed inside an autophagolysosome containing dense granular material. (I) A Caco-2 cell showing a clear apical membrane surface with microvilli. C. concisus UNSWCD which has maintained its spiral morphology is visualised inside an autophagolysosome.

Mentions: C. concisus UNSWCD cells found in close proximity to mitochondria were encapsulated by a fine layer (Fig. 7A, B), which could be indicative of early stage phagosome formation in Caco-2 cells. Filament arrangements were also identified near the bacterium (Fig. 7B, C). C. concisus UNSWCD were found to associate with autophagosomes showing a fine double membrane, one of their distinct characteristics (Fig. 7D). They were also found in close proximity to intermediary phagosomes (Fig. 7E), and in fusion with larger phagosomes containing both a vacuole compartment and an intermediary phagosome (Fig. 7F, G). Moreover, C. concisus UNSWCD can be found inside autophagolysosomes with dense granular material (Fig. 7H, I), in some cases its spiral morphology being still detectable (Fig. 7I).


The role of autophagy in the intracellular survival of Campylobacter concisus.

Burgos-Portugal JA, Mitchell HM, Castaño-Rodríguez N, Kaakoush NO - FEBS Open Bio (2014)

Visualization of the internalization of C. concisus into Caco-2 cells using transmission electron microscopy. (A) C. concisus UNSWCD invading inside Caco-2 cells shown in cross section measuring 0.5 μm wide. The bacterial cell appears to be in close proximity to a mitochondrion with a fine layer surrounding them. (B) A membrane forms around internalized C. concisus UNSWCD. A fine layer surrounds the bacterium alongside mitochondria and could be indicative of early stage phagosome formation in Caco-2 cells. (C) The appearance of fine filament arrangements close to the bacterium. (D) C. concisus UNSWCD associated with an autophagosome showing a double membrane. (E) C. concisus UNSWCD shown in cross section (0.5 μm wide) in close proximity to an intermediary phagosome. The bacterium appears to be within a vacuole. (F) C. concisus UNSWCD shown in cross section (0.5 μm wide). The bacterium is shown in close proximity with a vacuole compartment forming one large phagosome. (G) High magnification image of the bacterium fusing with a vacuole compartment. (H) C. concisus UNSWCD (0.5 μm wide) observed inside an autophagolysosome containing dense granular material. (I) A Caco-2 cell showing a clear apical membrane surface with microvilli. C. concisus UNSWCD which has maintained its spiral morphology is visualised inside an autophagolysosome.
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Related In: Results  -  Collection

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f0035: Visualization of the internalization of C. concisus into Caco-2 cells using transmission electron microscopy. (A) C. concisus UNSWCD invading inside Caco-2 cells shown in cross section measuring 0.5 μm wide. The bacterial cell appears to be in close proximity to a mitochondrion with a fine layer surrounding them. (B) A membrane forms around internalized C. concisus UNSWCD. A fine layer surrounds the bacterium alongside mitochondria and could be indicative of early stage phagosome formation in Caco-2 cells. (C) The appearance of fine filament arrangements close to the bacterium. (D) C. concisus UNSWCD associated with an autophagosome showing a double membrane. (E) C. concisus UNSWCD shown in cross section (0.5 μm wide) in close proximity to an intermediary phagosome. The bacterium appears to be within a vacuole. (F) C. concisus UNSWCD shown in cross section (0.5 μm wide). The bacterium is shown in close proximity with a vacuole compartment forming one large phagosome. (G) High magnification image of the bacterium fusing with a vacuole compartment. (H) C. concisus UNSWCD (0.5 μm wide) observed inside an autophagolysosome containing dense granular material. (I) A Caco-2 cell showing a clear apical membrane surface with microvilli. C. concisus UNSWCD which has maintained its spiral morphology is visualised inside an autophagolysosome.
Mentions: C. concisus UNSWCD cells found in close proximity to mitochondria were encapsulated by a fine layer (Fig. 7A, B), which could be indicative of early stage phagosome formation in Caco-2 cells. Filament arrangements were also identified near the bacterium (Fig. 7B, C). C. concisus UNSWCD were found to associate with autophagosomes showing a fine double membrane, one of their distinct characteristics (Fig. 7D). They were also found in close proximity to intermediary phagosomes (Fig. 7E), and in fusion with larger phagosomes containing both a vacuole compartment and an intermediary phagosome (Fig. 7F, G). Moreover, C. concisus UNSWCD can be found inside autophagolysosomes with dense granular material (Fig. 7H, I), in some cases its spiral morphology being still detectable (Fig. 7I).

Bottom Line: Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance.C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition.Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

View Article: PubMed Central - PubMed

Affiliation: School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, NSW 2052, Australia.

ABSTRACT
Campylobacter concisus is an emerging pathogen that has been associated with gastrointestinal diseases. Given the importance of autophagy for the elimination of intracellular bacteria and the subversion of this process by pathogenic bacteria, we investigated the role of autophagy in C. concisus intracellular survival. Gentamicin protection assays were employed to assess intracellular levels of C. concisus within Caco-2 cells, following autophagy induction and inhibition. To assess the interaction between C. concisus and autophagosomes, confocal microscopy, scanning electron microscopy, and transmission electron microscopy were employed. Expression levels of 84 genes involved in the autophagy process were measured using qPCR. Autophagy inhibition resulted in two- to four-fold increases in intracellular levels of C. concisus within Caco-2 cells, while autophagy induction resulted in a significant reduction in intracellular levels or bacterial clearance. C. concisus strains with low intracellular survival levels showed a dramatic increase in these levels upon autophagy inhibition. Confocal microscopy showed co-localization of the bacterium with autophagosomes, while transmission electron microscopy identified intracellular bacteria persisting within autophagic vesicles. Further, qPCR showed that following infection, 13 genes involved in the autophagy process were significantly regulated, and a further five showed borderline results, with an overall indication towards a dampening effect exerted by the bacterium on this process. Our data collectively indicates that while autophagy is important for the clearance of C. concisus, some strains may manipulate this process to benefit their intracellular survival.

No MeSH data available.


Related in: MedlinePlus