Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.

CagA protein contributes to pro-inflammatory responses during H. pylori infection, following its intracellular delivery to gastric epithelial cells. Here, we report for the first time in an isogenic background, on the subtle role of CagA phosphorylation on terminal EPIYA-C motifs in the transcriptio...

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Autores principales:	Konstantinos S Papadakos, Ioanna S Sougleri, Andreas F Mentis, Efstathios Hatziloukas, Dionyssios N Sgouras
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2013
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/340b7432385f491c87ea1bb462a7c2d0
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spelling	oai:doaj.org-article:340b7432385f491c87ea1bb462a7c2d02021-11-18T07:58:14ZPresence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.1932-620310.1371/journal.pone.0056291https://doaj.org/article/340b7432385f491c87ea1bb462a7c2d02013-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/23409168/?tool=EBIhttps://doaj.org/toc/1932-6203CagA protein contributes to pro-inflammatory responses during H. pylori infection, following its intracellular delivery to gastric epithelial cells. Here, we report for the first time in an isogenic background, on the subtle role of CagA phosphorylation on terminal EPIYA-C motifs in the transcriptional activation and expression of IL-8. We utilized isogenic H. pylori mutants of P12 reference strain, expressing CagA with varying number of EPIYA-C motifs and the corresponding phosphorylation defective EPIFA-C motifs while preserving intact the CM multimerization motifs. These mutants had been previously closely scrutinized in terms of type IV secretion system functionality, CagA translocation and its subsequent phosphorylation. Following infection of gastric epithelial cell lines, transcriptional activation of IL-8 gene and secreted IL-8 levels were found to be strictly dependent upon the functionality of the EPIYA-C phosphorylation motifs, as EPIFA-C phosphorylation-deficient CagA expression failed to induce full IL-8 transcriptional activity. Interestingly, levels of IL-8 gene activation and of secreted IL-8 were the same, irrespective of the number of EPIYA-C terminal repeats. We monitored IkBα phosphorylation and confirmed CagA involvement in NF-kB activation. Furthermore, we observed that presence of EPIYA-C functional phosphorylation motifs contributed to NF-kB activation. NF-kB upstream signaling events, such as early ERK1/2 and AKT activation were confirmed to be independent of EPIYA-C phosphorylation. On the contrary, use of TAK1 specific inhibitor 5Z-7-Oxozeaenol resulted in complete arrest of IL-8 secretion, in a dose-dependent manner, irrespective of CagA status. H. pylori-infected TAK1(-/-) mouse embryonic fibroblasts (MEFs) failed to induce NF-kB activity, unlike the respective control MEFs. CagA and TAK1 were found to immunoprecipitate together, irrespective of CagA EPIYA-C status, thus confirming earlier reports of TAK1 and CagA protein interaction. Our data suggest that CagA may potentially interfere with TAK1 activity during NF-kB activation for IL-8 induction in early H. pylori infection.Konstantinos S PapadakosIoanna S SougleriAndreas F MentisEfstathios HatziloukasDionyssios N SgourasPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 8, Iss 2, p e56291 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Konstantinos S Papadakos Ioanna S Sougleri Andreas F Mentis Efstathios Hatziloukas Dionyssios N Sgouras Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.
description	CagA protein contributes to pro-inflammatory responses during H. pylori infection, following its intracellular delivery to gastric epithelial cells. Here, we report for the first time in an isogenic background, on the subtle role of CagA phosphorylation on terminal EPIYA-C motifs in the transcriptional activation and expression of IL-8. We utilized isogenic H. pylori mutants of P12 reference strain, expressing CagA with varying number of EPIYA-C motifs and the corresponding phosphorylation defective EPIFA-C motifs while preserving intact the CM multimerization motifs. These mutants had been previously closely scrutinized in terms of type IV secretion system functionality, CagA translocation and its subsequent phosphorylation. Following infection of gastric epithelial cell lines, transcriptional activation of IL-8 gene and secreted IL-8 levels were found to be strictly dependent upon the functionality of the EPIYA-C phosphorylation motifs, as EPIFA-C phosphorylation-deficient CagA expression failed to induce full IL-8 transcriptional activity. Interestingly, levels of IL-8 gene activation and of secreted IL-8 were the same, irrespective of the number of EPIYA-C terminal repeats. We monitored IkBα phosphorylation and confirmed CagA involvement in NF-kB activation. Furthermore, we observed that presence of EPIYA-C functional phosphorylation motifs contributed to NF-kB activation. NF-kB upstream signaling events, such as early ERK1/2 and AKT activation were confirmed to be independent of EPIYA-C phosphorylation. On the contrary, use of TAK1 specific inhibitor 5Z-7-Oxozeaenol resulted in complete arrest of IL-8 secretion, in a dose-dependent manner, irrespective of CagA status. H. pylori-infected TAK1(-/-) mouse embryonic fibroblasts (MEFs) failed to induce NF-kB activity, unlike the respective control MEFs. CagA and TAK1 were found to immunoprecipitate together, irrespective of CagA EPIYA-C status, thus confirming earlier reports of TAK1 and CagA protein interaction. Our data suggest that CagA may potentially interfere with TAK1 activity during NF-kB activation for IL-8 induction in early H. pylori infection.
format	article
author	Konstantinos S Papadakos Ioanna S Sougleri Andreas F Mentis Efstathios Hatziloukas Dionyssios N Sgouras
author_facet	Konstantinos S Papadakos Ioanna S Sougleri Andreas F Mentis Efstathios Hatziloukas Dionyssios N Sgouras
author_sort	Konstantinos S Papadakos
title	Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.
title_short	Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.
title_full	Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.
title_fullStr	Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.
title_full_unstemmed	Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.
title_sort	presence of terminal epiya phosphorylation motifs in helicobacter pylori caga contributes to il-8 secretion, irrespective of the number of repeats.
publisher	Public Library of Science (PLoS)
publishDate	2013
url	https://doaj.org/article/340b7432385f491c87ea1bb462a7c2d0
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Presence of terminal EPIYA phosphorylation motifs in Helicobacter pylori CagA contributes to IL-8 secretion, irrespective of the number of repeats.

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