Molecular mechanisms of virulence gene regulation in gastric cell adhered helicobacter pylori and development of therapeutic nano-particles
IR@IICB: CSIR-Indian Institute of Chemical Biology, Kolkata
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| Title |
Molecular mechanisms of virulence gene regulation in gastric cell adhered helicobacter pylori and development of therapeutic nano-particles
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| Creator |
Bhattacharya, Saurabh
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| Subject |
Infectious Diseases and Immunology
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| Description |
Helicobacter pylori is a major human pathogen and is associated with chronic gastric inflammation, peptic ulcer disease and gastric cancer. In this study, it was observed
that adherence of H. pylori to the gastric epithelial cell line AGS strongly induces expression of the genes HP0102, fliK and cagA. HP0102 was identified to be involved
in the glycosylation of bacterial lipopolysachharides (LPS), whereas fliK is known to be required for flagellar biogenesis. One of the striking features of gastric epithelial cells (AGS) infected with ΔHP0102 and ΔfliK strains is their reduced “hummingbird” appearance and IL-8 secretion, a phenomenon attributed to the expression and translocation of the bacterial CagA protein, a major virulence factor. Interestingly, upregulation of cagA expression was not observed in AGS adhered H.pylori ΔHP0102 and ΔfliK mutants. Subsequently, it was identified that both HP0102 and fliK independently influence the expression of the alternative factor 28. Induction of fliA, encoding 28 was also impaired in the mutants following
adherence. Furthermore, it was identified that direct binding of σ28-RNAP initiated transcription of cagA from an alternative downstream promoter active specifically in
AGS-adhered wild type H. pylori but not in ΔHP0102 and ΔfliK strains. Thus, in this study we have identified that HP0102 and FliK are involved in the regulation of
cagA expression in host cell adhered H. pylori. Further investigation suggested that the chemotaxis and the acid escape response of the ΔHP0102 strain was affected, phenotypes that might account for its previously
reported deficiency in mice colonization. Finally, proteome analysis suggested that HP0102 is not only involved in the regulation of cagA but also in the fine tuning of
the expression of several other genes. A small non-coding RNA, HPnc2630 was also one of the genes strongly upregulated following adherence of H. pylori to AGS cells. Transcriptome analysis revealed that HPnc2630 regulates the expression of several genes, thereby influencing the
pathogenesis of the bacteria. Affinity capture studies suggested that this sRNA interacts with specific proteins CytC peroxidase & γ-glutamyl transpeptidase (GGT)
and probably sequesters them. GGT is a known positive modulator of VacA activity. Interestingly, a ΔsRNA mutant strain exhibited increased ability to adhere to AGS
cells and induced more vacuolation in these cells, thereby confirming the role of the sRNA in the regulation of VacA.
In the final part of the study, Polyethyleneinime (PEI) capped ZnO nano-particles (NPs) have been developed as a putative therapeutic. These have better dispersion,
stability under low pH conditions and are more potent when compared to the conventional ZnO NPs in their ability to inhibit the growth of antibiotic resistant H. pylori. Furthermore, they internalize into the bacterial cells, generate huge amounts of reactive oxygen species (ROS), leading to membrane damage and morphological
transition from the rod to coccoid form along with concomitant degradation of stable RNA molecules in these cells. Toxicity studies suggest that a dose < 25 μg/ml
is safe for use in human cells. At this concentration, ZnO-PEI NPs show notable synergy in combination with antibiotics to inhibit H. pylori
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| Date |
2016-02-25
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| Type |
Thesis
NonPeerReviewed |
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| Format |
application/pdf
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| Identifier |
http://www.eprints.iicb.res.in/2502/1/THESIS_SAURABH_BHATTACHARYA.pdf
Bhattacharya, Saurabh (2016) Molecular mechanisms of virulence gene regulation in gastric cell adhered helicobacter pylori and development of therapeutic nano-particles. PhD thesis, J U. |
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| Relation |
http://www.eprints.iicb.res.in/2502/
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