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Functional plasticity in the type IV secretion system of Helicobacter pylori.

Barrozo RM, Cooke CL, Hansen LM, Lam AM, Gaddy JA, Johnson EM, Cariaga TA, Suarez G, Peek RM, Cover TL, Solnick JV - PLoS Pathog. (2013)

Bottom Line: CagY is an essential component of the H. pylori T4SS that has an unusual sequence structure, in which an extraordinary number of direct DNA repeats is predicted to cause rearrangements that invariably yield in-frame insertions or deletions.Here we demonstrate in murine and non-human primate models that immune-driven host selection of rearrangements in CagY is sufficient to cause gain or loss of function in the H. pylori T4SS.We propose that CagY functions as a sort of molecular switch or perhaps a rheostat that alters the function of the T4SS and "tunes" the host inflammatory response so as to maximize persistent infection.

View Article: PubMed Central - PubMed

Affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.

ABSTRACT
Helicobacter pylori causes clinical disease primarily in those individuals infected with a strain that carries the cytotoxin associated gene pathogenicity island (cagPAI). The cagPAI encodes a type IV secretion system (T4SS) that injects the CagA oncoprotein into epithelial cells and is required for induction of the pro-inflammatory cytokine, interleukin-8 (IL-8). CagY is an essential component of the H. pylori T4SS that has an unusual sequence structure, in which an extraordinary number of direct DNA repeats is predicted to cause rearrangements that invariably yield in-frame insertions or deletions. Here we demonstrate in murine and non-human primate models that immune-driven host selection of rearrangements in CagY is sufficient to cause gain or loss of function in the H. pylori T4SS. We propose that CagY functions as a sort of molecular switch or perhaps a rheostat that alters the function of the T4SS and "tunes" the host inflammatory response so as to maximize persistent infection.

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Mouse adapted H. pylori strain SS1 expresses a CagY that is not functional for induction of IL-8 or translocation of CagA.(A) H. pylori was isolated from C57BL/6 WT or RAG1−/− mice (N = 3–6/time point) 8 weeks after experimental infection with H. pylori PMSS1. Individual colonies (3–6/mouse) were co-cultured with AGS cells, and ELISA was used to measure IL-8 levels, which were normalized to the PMSS1 positive control (line = mean). Each data point represents the results from a single colony. Induction of IL-8 in colonies isolated from WT mice was significantly lower than in RAG1−/− mice, and was associated with changes in cagY PCR-RFLP (open circles). (B) cagY in H. pylori strain SS1 is larger than that in the progenitor strain PMSS1, and has a different fingerprint on PCR-RFLP. (C) Deletion of cagY from WT H. pylori PMSS1 reduced the induction of IL-8 and eliminated translocation of CagA, which were recovered when the WT PMSS1 cagY gene was restored (▵Y[PMSS1]. However, replacement of the PMSS1 cagY gene with that from H. pylori SS1 (▵Y [SS1]) showed reduced levels of IL-8 and no CagA translocation. (D) WT H. pylori SS1 showed little induction of IL-8 and no CagA translocation, and it was unaffected by deletion of cagY or restoration of the WT SS1 cagY allele. However, replacement of the WT SS1 cagY allele with that from PMSS1 markedly increased IL-8 induction and CagA translocation, though not to the level of PMSS1. All assays represent the mean ±SEM of 3 replicates. **P<0.01; ***P<0.001.
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ppat-1003189-g009: Mouse adapted H. pylori strain SS1 expresses a CagY that is not functional for induction of IL-8 or translocation of CagA.(A) H. pylori was isolated from C57BL/6 WT or RAG1−/− mice (N = 3–6/time point) 8 weeks after experimental infection with H. pylori PMSS1. Individual colonies (3–6/mouse) were co-cultured with AGS cells, and ELISA was used to measure IL-8 levels, which were normalized to the PMSS1 positive control (line = mean). Each data point represents the results from a single colony. Induction of IL-8 in colonies isolated from WT mice was significantly lower than in RAG1−/− mice, and was associated with changes in cagY PCR-RFLP (open circles). (B) cagY in H. pylori strain SS1 is larger than that in the progenitor strain PMSS1, and has a different fingerprint on PCR-RFLP. (C) Deletion of cagY from WT H. pylori PMSS1 reduced the induction of IL-8 and eliminated translocation of CagA, which were recovered when the WT PMSS1 cagY gene was restored (▵Y[PMSS1]. However, replacement of the PMSS1 cagY gene with that from H. pylori SS1 (▵Y [SS1]) showed reduced levels of IL-8 and no CagA translocation. (D) WT H. pylori SS1 showed little induction of IL-8 and no CagA translocation, and it was unaffected by deletion of cagY or restoration of the WT SS1 cagY allele. However, replacement of the WT SS1 cagY allele with that from PMSS1 markedly increased IL-8 induction and CagA translocation, though not to the level of PMSS1. All assays represent the mean ±SEM of 3 replicates. **P<0.01; ***P<0.001.

Mentions: Studies of H. pylori pathogenesis were long hampered by the inability of investigators to successfully colonize mice. Since the difficulty was attributed primarily to H. pylori strain differences, mouse-adapted strain SS1 was derived, which has become the standard for animal experimentation [33]. However, it was later realized that H. pylori SS1 did not induce IL-8 or translocate CagA [20], [21], despite having an intact cagPAI detected by microarray [34]. The reason for this is unknown. It was recently reported that the original human isolate, designated pre-mouse SS1 (PMSS1), does have a functional cagPAI [35]. We therefore hypothesized that SS1 had undergone recombination in cagY during mouse passage that eliminated its capacity to induce IL-8 and translocate CagA. To test this hypothesis, we first inoculated PMSS1 into WT C57BL/6 and RAG1−/− mice, and examined IL-8 induction and cagY RFLP in colonies recovered 8 weeks PI. Similar to the results with strain J166 (Figure 3), colonies from WT but not RAG1−/− mice showed loss of IL-8 induction that was associated with recombination in cagY (Figure 9A). These results are consistent with a previous report demonstrating loss of T4SS function after challenge with PMSS1 in adult but not neonatal mice, which control effector T cell responses by H. pylori-specific regulatory T cells [35].


Functional plasticity in the type IV secretion system of Helicobacter pylori.

Barrozo RM, Cooke CL, Hansen LM, Lam AM, Gaddy JA, Johnson EM, Cariaga TA, Suarez G, Peek RM, Cover TL, Solnick JV - PLoS Pathog. (2013)

Mouse adapted H. pylori strain SS1 expresses a CagY that is not functional for induction of IL-8 or translocation of CagA.(A) H. pylori was isolated from C57BL/6 WT or RAG1−/− mice (N = 3–6/time point) 8 weeks after experimental infection with H. pylori PMSS1. Individual colonies (3–6/mouse) were co-cultured with AGS cells, and ELISA was used to measure IL-8 levels, which were normalized to the PMSS1 positive control (line = mean). Each data point represents the results from a single colony. Induction of IL-8 in colonies isolated from WT mice was significantly lower than in RAG1−/− mice, and was associated with changes in cagY PCR-RFLP (open circles). (B) cagY in H. pylori strain SS1 is larger than that in the progenitor strain PMSS1, and has a different fingerprint on PCR-RFLP. (C) Deletion of cagY from WT H. pylori PMSS1 reduced the induction of IL-8 and eliminated translocation of CagA, which were recovered when the WT PMSS1 cagY gene was restored (▵Y[PMSS1]. However, replacement of the PMSS1 cagY gene with that from H. pylori SS1 (▵Y [SS1]) showed reduced levels of IL-8 and no CagA translocation. (D) WT H. pylori SS1 showed little induction of IL-8 and no CagA translocation, and it was unaffected by deletion of cagY or restoration of the WT SS1 cagY allele. However, replacement of the WT SS1 cagY allele with that from PMSS1 markedly increased IL-8 induction and CagA translocation, though not to the level of PMSS1. All assays represent the mean ±SEM of 3 replicates. **P<0.01; ***P<0.001.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3585145&req=5

ppat-1003189-g009: Mouse adapted H. pylori strain SS1 expresses a CagY that is not functional for induction of IL-8 or translocation of CagA.(A) H. pylori was isolated from C57BL/6 WT or RAG1−/− mice (N = 3–6/time point) 8 weeks after experimental infection with H. pylori PMSS1. Individual colonies (3–6/mouse) were co-cultured with AGS cells, and ELISA was used to measure IL-8 levels, which were normalized to the PMSS1 positive control (line = mean). Each data point represents the results from a single colony. Induction of IL-8 in colonies isolated from WT mice was significantly lower than in RAG1−/− mice, and was associated with changes in cagY PCR-RFLP (open circles). (B) cagY in H. pylori strain SS1 is larger than that in the progenitor strain PMSS1, and has a different fingerprint on PCR-RFLP. (C) Deletion of cagY from WT H. pylori PMSS1 reduced the induction of IL-8 and eliminated translocation of CagA, which were recovered when the WT PMSS1 cagY gene was restored (▵Y[PMSS1]. However, replacement of the PMSS1 cagY gene with that from H. pylori SS1 (▵Y [SS1]) showed reduced levels of IL-8 and no CagA translocation. (D) WT H. pylori SS1 showed little induction of IL-8 and no CagA translocation, and it was unaffected by deletion of cagY or restoration of the WT SS1 cagY allele. However, replacement of the WT SS1 cagY allele with that from PMSS1 markedly increased IL-8 induction and CagA translocation, though not to the level of PMSS1. All assays represent the mean ±SEM of 3 replicates. **P<0.01; ***P<0.001.
Mentions: Studies of H. pylori pathogenesis were long hampered by the inability of investigators to successfully colonize mice. Since the difficulty was attributed primarily to H. pylori strain differences, mouse-adapted strain SS1 was derived, which has become the standard for animal experimentation [33]. However, it was later realized that H. pylori SS1 did not induce IL-8 or translocate CagA [20], [21], despite having an intact cagPAI detected by microarray [34]. The reason for this is unknown. It was recently reported that the original human isolate, designated pre-mouse SS1 (PMSS1), does have a functional cagPAI [35]. We therefore hypothesized that SS1 had undergone recombination in cagY during mouse passage that eliminated its capacity to induce IL-8 and translocate CagA. To test this hypothesis, we first inoculated PMSS1 into WT C57BL/6 and RAG1−/− mice, and examined IL-8 induction and cagY RFLP in colonies recovered 8 weeks PI. Similar to the results with strain J166 (Figure 3), colonies from WT but not RAG1−/− mice showed loss of IL-8 induction that was associated with recombination in cagY (Figure 9A). These results are consistent with a previous report demonstrating loss of T4SS function after challenge with PMSS1 in adult but not neonatal mice, which control effector T cell responses by H. pylori-specific regulatory T cells [35].

Bottom Line: CagY is an essential component of the H. pylori T4SS that has an unusual sequence structure, in which an extraordinary number of direct DNA repeats is predicted to cause rearrangements that invariably yield in-frame insertions or deletions.Here we demonstrate in murine and non-human primate models that immune-driven host selection of rearrangements in CagY is sufficient to cause gain or loss of function in the H. pylori T4SS.We propose that CagY functions as a sort of molecular switch or perhaps a rheostat that alters the function of the T4SS and "tunes" the host inflammatory response so as to maximize persistent infection.

View Article: PubMed Central - PubMed

Affiliation: Center for Comparative Medicine, University of California Davis, Davis, California, United States of America.

ABSTRACT
Helicobacter pylori causes clinical disease primarily in those individuals infected with a strain that carries the cytotoxin associated gene pathogenicity island (cagPAI). The cagPAI encodes a type IV secretion system (T4SS) that injects the CagA oncoprotein into epithelial cells and is required for induction of the pro-inflammatory cytokine, interleukin-8 (IL-8). CagY is an essential component of the H. pylori T4SS that has an unusual sequence structure, in which an extraordinary number of direct DNA repeats is predicted to cause rearrangements that invariably yield in-frame insertions or deletions. Here we demonstrate in murine and non-human primate models that immune-driven host selection of rearrangements in CagY is sufficient to cause gain or loss of function in the H. pylori T4SS. We propose that CagY functions as a sort of molecular switch or perhaps a rheostat that alters the function of the T4SS and "tunes" the host inflammatory response so as to maximize persistent infection.

Show MeSH
Related in: MedlinePlus