Abstract
Helicobacter pylori (Hp) CagA protein has a key role in the development of gastric cancer by the intruding in many intracellular processes of host human cell. Endoplasmic reticulum (ER) stress is an essential process for cellular homeostasis that modulates survival and death and is linked to several complex diseases including cancer. CagA protein is found in the serum of Hp-positive individuals and also in the supernatant of Hp culture. Limited studies report that recombinant CagA can alter gene expression and signaling pathways and induce the death of human cells. In this study, we investigated the effect of exogenous recombinant CagA protein treatment on ER stress and autophagy of human cell. AGS, MKN45, and HEK293 cells were treated with 1 µg/ml of recombinant CagA protein and then ER stress was studied by quantitative-PCR of spliced XBP-1 mRNA, immunofluorescence staining of ATF6 protein nuclear localization and real-time quantitative-PCR and/or western blot expression of GRP78, GRP94, ATF4 and CHOP genes. Autophagy was studied by western blot assessment of the conversion of LC3-I to LC3-II and LC3 aggregation. Cell proliferation and death were investigated by MTT assay and trypan blue staining respectively. As result, treatment with recombinant CagA enhanced XBP-1mRNA splicing, nuclear localization of ATF6, and the expression of ER stress signaling target genes in the cells. Recombinant CagA also induced LC3 protein conversion and aggregation in the cells. Reduced cell proliferation and increased cell death were determined in the cells treated with recombinant CagA. These results show that exogenous recombinant CagA protein causes cell death by inducing ER stress and autophagy in human cells. We conclude that CagA protein exogenously localizes in/on human cells and induces ER stress via disturbing protein machinery leading the human cell to death, however, the mechanism of CagA-host cell interaction is to be investigated.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
The authors welcome journals to consider this article for peer-review and publication.
Abbreviations
- ANOVA
- Analysis of variance
- ATF6
- Activating transcription factor 6
- bZIP
- Basic leucine zipper
- C2Cer
- C2-ceramide
- CagA
- Cytotoxin associated gene A
- cagPAI
- Cag pathogenicity island
- DMSO
- Dimethyl sulfoxide
- ER
- Endoplasmic reticulum
- ERAD
- Endoplasmic reticulum-associated degradation
- Hp
- Helicobacter pylori
- HRP
- Horseradish peroxidase
- IDT
- Integrated DNA Technology
- IRE1α
- Inositol-requiring kinase 1α
- PBS
- Phosphate buffered saline
- PERK
- PRKR-like ER kinase
- T4SS
- Type IV secretion system
- Tm
- Tunicamycin
- UT
- Untreated
- XBP-1
- X-box-binding protein.