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Gadd45α activity is the principal effector of Shigella mitochondria-dependent epithelial cell death in vitro and ex vivo.

Lembo-Fazio L, Nigro G, Noël G, Rossi G, Chiara F, Tsilingiri K, Rescigno M, Rasola A, Bernardini ML - Cell Death Dis (2011)

Bottom Line: Modulation of death is a pathogen strategy to establish residence and promote survival in host cells and tissues.However, they are equally able to protect host cells from death.To explore these aspects, we have exploited both, the HeLa cell invasion assay and a novel ex vivo human colon organ culture model of infection that mimics natural conditions of shigellosis.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Biologia e Biotecnologie Charles Darwin, Sapienza-Università di Roma, Roma, Italy.

ABSTRACT
Modulation of death is a pathogen strategy to establish residence and promote survival in host cells and tissues. Shigella spp. are human pathogens that invade colonic mucosa, where they provoke lesions caused by their ability to manipulate the host cell responses. Shigella spp. induce various types of cell death in different cell populations. However, they are equally able to protect host cells from death. Here, we have investigated on the molecular mechanisms and cell effectors governing the balance between survival and death in epithelial cells infected with Shigella. To explore these aspects, we have exploited both, the HeLa cell invasion assay and a novel ex vivo human colon organ culture model of infection that mimics natural conditions of shigellosis. Our results definitely show that Shigella induces a rapid intrinsic apoptosis of infected cells, via mitochondrial depolarization and the ensuing caspase-9 activation. Moreover, for the first time we identify the eukaryotic stress-response factor growth arrest and DNA damage 45α as a key player in the induction of the apoptotic process elicited by Shigella in epithelial cells, revealing an unexplored role of this molecule in the course of infections sustained by invasive pathogens.

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Related in: MedlinePlus

Modulation of pro- and anti-apoptotic gene expression in HeLa cells on infection with S. flexneri M90T. (a) Transcriptional profile of pro- and anti-apoptotic genes expressed by HeLa cells infected with S. flexneri M90T (MOI 100) at 1, 3 and 5 h of incubation p.i. Red and green colors represent up- and downregulation of gene expression, respectively, as compared with uninfected cells. The key for intensity of expression is indicated under the bar. (b) Validation/determination through qPCR for a representative subset of genes whose expression is highly modulated by Shigella infection. Results are normalized to the internal gapdh gene control and are presented on a logarithmic scale as the ratio of gene expression between infected and uninfected HeLa cells. (c and d) Western immunoblot analysis of changes in protein expression levels of HeLa cells following Shigella infection. Cell lysis was carried out at the reported time points. Blots were probed with actin as a loading control
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fig4: Modulation of pro- and anti-apoptotic gene expression in HeLa cells on infection with S. flexneri M90T. (a) Transcriptional profile of pro- and anti-apoptotic genes expressed by HeLa cells infected with S. flexneri M90T (MOI 100) at 1, 3 and 5 h of incubation p.i. Red and green colors represent up- and downregulation of gene expression, respectively, as compared with uninfected cells. The key for intensity of expression is indicated under the bar. (b) Validation/determination through qPCR for a representative subset of genes whose expression is highly modulated by Shigella infection. Results are normalized to the internal gapdh gene control and are presented on a logarithmic scale as the ratio of gene expression between infected and uninfected HeLa cells. (c and d) Western immunoblot analysis of changes in protein expression levels of HeLa cells following Shigella infection. Cell lysis was carried out at the reported time points. Blots were probed with actin as a loading control

Mentions: We used an oligonucleotide array spotted with 135 apoptotic and anti-apoptotic genes (see Materials and methods section) and identified a panel of genes differentially expressed, which were subjected to hierarchical clustering to reveal expression trends (Figure 4a and Supplementary Table S1).


Gadd45α activity is the principal effector of Shigella mitochondria-dependent epithelial cell death in vitro and ex vivo.

Lembo-Fazio L, Nigro G, Noël G, Rossi G, Chiara F, Tsilingiri K, Rescigno M, Rasola A, Bernardini ML - Cell Death Dis (2011)

Modulation of pro- and anti-apoptotic gene expression in HeLa cells on infection with S. flexneri M90T. (a) Transcriptional profile of pro- and anti-apoptotic genes expressed by HeLa cells infected with S. flexneri M90T (MOI 100) at 1, 3 and 5 h of incubation p.i. Red and green colors represent up- and downregulation of gene expression, respectively, as compared with uninfected cells. The key for intensity of expression is indicated under the bar. (b) Validation/determination through qPCR for a representative subset of genes whose expression is highly modulated by Shigella infection. Results are normalized to the internal gapdh gene control and are presented on a logarithmic scale as the ratio of gene expression between infected and uninfected HeLa cells. (c and d) Western immunoblot analysis of changes in protein expression levels of HeLa cells following Shigella infection. Cell lysis was carried out at the reported time points. Blots were probed with actin as a loading control
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3101704&req=5

fig4: Modulation of pro- and anti-apoptotic gene expression in HeLa cells on infection with S. flexneri M90T. (a) Transcriptional profile of pro- and anti-apoptotic genes expressed by HeLa cells infected with S. flexneri M90T (MOI 100) at 1, 3 and 5 h of incubation p.i. Red and green colors represent up- and downregulation of gene expression, respectively, as compared with uninfected cells. The key for intensity of expression is indicated under the bar. (b) Validation/determination through qPCR for a representative subset of genes whose expression is highly modulated by Shigella infection. Results are normalized to the internal gapdh gene control and are presented on a logarithmic scale as the ratio of gene expression between infected and uninfected HeLa cells. (c and d) Western immunoblot analysis of changes in protein expression levels of HeLa cells following Shigella infection. Cell lysis was carried out at the reported time points. Blots were probed with actin as a loading control
Mentions: We used an oligonucleotide array spotted with 135 apoptotic and anti-apoptotic genes (see Materials and methods section) and identified a panel of genes differentially expressed, which were subjected to hierarchical clustering to reveal expression trends (Figure 4a and Supplementary Table S1).

Bottom Line: Modulation of death is a pathogen strategy to establish residence and promote survival in host cells and tissues.However, they are equally able to protect host cells from death.To explore these aspects, we have exploited both, the HeLa cell invasion assay and a novel ex vivo human colon organ culture model of infection that mimics natural conditions of shigellosis.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Biologia e Biotecnologie Charles Darwin, Sapienza-Università di Roma, Roma, Italy.

ABSTRACT
Modulation of death is a pathogen strategy to establish residence and promote survival in host cells and tissues. Shigella spp. are human pathogens that invade colonic mucosa, where they provoke lesions caused by their ability to manipulate the host cell responses. Shigella spp. induce various types of cell death in different cell populations. However, they are equally able to protect host cells from death. Here, we have investigated on the molecular mechanisms and cell effectors governing the balance between survival and death in epithelial cells infected with Shigella. To explore these aspects, we have exploited both, the HeLa cell invasion assay and a novel ex vivo human colon organ culture model of infection that mimics natural conditions of shigellosis. Our results definitely show that Shigella induces a rapid intrinsic apoptosis of infected cells, via mitochondrial depolarization and the ensuing caspase-9 activation. Moreover, for the first time we identify the eukaryotic stress-response factor growth arrest and DNA damage 45α as a key player in the induction of the apoptotic process elicited by Shigella in epithelial cells, revealing an unexplored role of this molecule in the course of infections sustained by invasive pathogens.

Show MeSH
Related in: MedlinePlus