Limits...
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.

Show MeSH

Related in: MedlinePlus

Loss of the capacity to induce IL-8 and change cagY during infection of mice is dependent on an intact host immune system.H. pylori was isolated from C57BL/6 WT (A) or RAG2−/− (B) mice (N = 3–6/time point) up to 16 weeks after experimental infection with H. pylori WT J166. 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 WT J166 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 RAG2−/− mice at 12 and 16 weeks PI (P<0.01). Changes in cagY (open circles) were detected by PCR-RFLP in 28 of 70 colonies from WT mice but in 0 of 64 colonies from RAG2−/− mice (Fishers exact test, P<0.0001). Output strains from WT C57BL/6 mice were analyzed by cagY PCR-RFLP and compared to WT H. pylori J166 (dark blue) and to one another (C). Each pie chart represents the unique cagY RFLP patterns identified in a single mouse from 2 to 16 weeks PI, and is positioned according to the mean IL-8 induction by colonies recovered from that mouse.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3585145&req=5

ppat-1003189-g003: Loss of the capacity to induce IL-8 and change cagY during infection of mice is dependent on an intact host immune system.H. pylori was isolated from C57BL/6 WT (A) or RAG2−/− (B) mice (N = 3–6/time point) up to 16 weeks after experimental infection with H. pylori WT J166. 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 WT J166 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 RAG2−/− mice at 12 and 16 weeks PI (P<0.01). Changes in cagY (open circles) were detected by PCR-RFLP in 28 of 70 colonies from WT mice but in 0 of 64 colonies from RAG2−/− mice (Fishers exact test, P<0.0001). Output strains from WT C57BL/6 mice were analyzed by cagY PCR-RFLP and compared to WT H. pylori J166 (dark blue) and to one another (C). Each pie chart represents the unique cagY RFLP patterns identified in a single mouse from 2 to 16 weeks PI, and is positioned according to the mean IL-8 induction by colonies recovered from that mouse.

Mentions: Identification of the direct repeat structure of cagY suggested that frequent in-frame recombination events may be a mechanism of antigenic variation to avoid the host adaptive immune response [28]. To test this hypothesis, we inoculated WT H. pylori J166 into WT C57BL/6 and RAG2−/− mice, which do not have functional B or T cells and develop little or no gastric inflammation after H. pylori infection [29]. H. pylori colonization levels were approximately 10-fold higher in RAG2−/− mice compared to WT mice (Figure S5A). Similar to the results in rhesus monkeys, bacteria recovered from WT mice resembled input H. pylori early after challenge (Figure 3A). However, at 12 and 16 weeks PI, bacteria from WT mice showed a significant loss in IL-8 induction (P<0.01) and change in cagY (P<0.001) compared to colonies from RAG2−/− mice, which uniformly resembled WT J166 in IL-8 induction and showed no changes in cagY by RFLP analysis (Figure 3B). We next replaced the cagY allele in WT H. pylori J166 with that from mouse output strains that changed cagY and either lost (mOut1 and mOut2) or maintained (mOut3 and mOut4) IL-8 induction in AGS cells, which was confirmed in KATO III cells (Figure S1B). Similar to the results with rhesus output strains (Figure 2), induction of IL-8 and phosphorylation of CagA in mouse output strains were phenocopied when their cagY allele was used to replace that in WT J166 (Figure 4). Interestingly, the bacterial population within each individual mouse was relatively homogenous, showing either WT levels of IL-8 and cagY indistinguishable from input, or low IL-8 and one or at most two unique cagY variants (Figure 3C). These experiments demonstrate that CagY-mediated change in function of the H. pylori T4SS is dependent on an intact host immune system.


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)

Loss of the capacity to induce IL-8 and change cagY during infection of mice is dependent on an intact host immune system.H. pylori was isolated from C57BL/6 WT (A) or RAG2−/− (B) mice (N = 3–6/time point) up to 16 weeks after experimental infection with H. pylori WT J166. 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 WT J166 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 RAG2−/− mice at 12 and 16 weeks PI (P<0.01). Changes in cagY (open circles) were detected by PCR-RFLP in 28 of 70 colonies from WT mice but in 0 of 64 colonies from RAG2−/− mice (Fishers exact test, P<0.0001). Output strains from WT C57BL/6 mice were analyzed by cagY PCR-RFLP and compared to WT H. pylori J166 (dark blue) and to one another (C). Each pie chart represents the unique cagY RFLP patterns identified in a single mouse from 2 to 16 weeks PI, and is positioned according to the mean IL-8 induction by colonies recovered from that mouse.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585145&req=5

ppat-1003189-g003: Loss of the capacity to induce IL-8 and change cagY during infection of mice is dependent on an intact host immune system.H. pylori was isolated from C57BL/6 WT (A) or RAG2−/− (B) mice (N = 3–6/time point) up to 16 weeks after experimental infection with H. pylori WT J166. 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 WT J166 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 RAG2−/− mice at 12 and 16 weeks PI (P<0.01). Changes in cagY (open circles) were detected by PCR-RFLP in 28 of 70 colonies from WT mice but in 0 of 64 colonies from RAG2−/− mice (Fishers exact test, P<0.0001). Output strains from WT C57BL/6 mice were analyzed by cagY PCR-RFLP and compared to WT H. pylori J166 (dark blue) and to one another (C). Each pie chart represents the unique cagY RFLP patterns identified in a single mouse from 2 to 16 weeks PI, and is positioned according to the mean IL-8 induction by colonies recovered from that mouse.
Mentions: Identification of the direct repeat structure of cagY suggested that frequent in-frame recombination events may be a mechanism of antigenic variation to avoid the host adaptive immune response [28]. To test this hypothesis, we inoculated WT H. pylori J166 into WT C57BL/6 and RAG2−/− mice, which do not have functional B or T cells and develop little or no gastric inflammation after H. pylori infection [29]. H. pylori colonization levels were approximately 10-fold higher in RAG2−/− mice compared to WT mice (Figure S5A). Similar to the results in rhesus monkeys, bacteria recovered from WT mice resembled input H. pylori early after challenge (Figure 3A). However, at 12 and 16 weeks PI, bacteria from WT mice showed a significant loss in IL-8 induction (P<0.01) and change in cagY (P<0.001) compared to colonies from RAG2−/− mice, which uniformly resembled WT J166 in IL-8 induction and showed no changes in cagY by RFLP analysis (Figure 3B). We next replaced the cagY allele in WT H. pylori J166 with that from mouse output strains that changed cagY and either lost (mOut1 and mOut2) or maintained (mOut3 and mOut4) IL-8 induction in AGS cells, which was confirmed in KATO III cells (Figure S1B). Similar to the results with rhesus output strains (Figure 2), induction of IL-8 and phosphorylation of CagA in mouse output strains were phenocopied when their cagY allele was used to replace that in WT J166 (Figure 4). Interestingly, the bacterial population within each individual mouse was relatively homogenous, showing either WT levels of IL-8 and cagY indistinguishable from input, or low IL-8 and one or at most two unique cagY variants (Figure 3C). These experiments demonstrate that CagY-mediated change in function of the H. pylori T4SS is dependent on an intact host immune system.

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