A nonstructural 2B protein of enterovirus A71 increases cytosolic Ca2+ and induces apoptosis in human neuroblastoma SH-SY5Y cells
Enterovirus A71 (EV-A71) is one of the causative agents causing the hand-foot-mouth disease which associated with fatal neurological complications. Several sporadic outbreaks of EV-A71 infections have been recently reported from Asia-Pacific regions and potentially established endemicity in the area. Currently, there is no effective vaccine or antiviral drug for EV-A71 available. This may be attributable to the limited information about its pathogenesis. In this study, the recombinant nonstructural 2B protein of EV-A71 was successfully produced in human neuroblastoma SH-SY5Y cells and evaluated for its effects on induction of the cell apoptosis and the pathway involved. The EV-A71 2B-transfected SH-SY5Y cells showed significantly higher difference in the cell growth inhibition than the mock and the irrelevant protein controls. The transfected SH-SY5Y cells underwent apoptosis and showed the significant upregulation of caspase-9 (CASP9) and caspase-12 (CASP12) genes at 3- and 24-h post-transfection, respectively. Interestingly, the level of cytosolic Ca2+ was significantly elevated in the transfected SH-SY5Y cells at 6- and 12-h post-transfection. The caspase-9 is activated by mitochondrial signaling pathway while the caspase-12 is activated by ER signaling pathway. The results suggested that EV-A71 2B protein triggered transient increase of the cytosolic Ca2+ level and associated with ER-mitochondrial interactions that drive the caspase-dependent apoptosis pathways. The detailed mechanisms warrant further studies for understanding the implication of EV-A71 infection in neuropathogenesis. The gained knowledge is essential for the development of the effective therapeutics and antiviral drugs.
KeywordsNonstructural 2B protein Enterovirus A71 Human neuroblastoma cell Apoptosis Calcium signaling pathway
OS conducted the overall study and data analysis; PT supervised OS on flow cytometry experiments and conducted the data analysis; PS designed the experiments and conducted the data analysis; PR constructed the recombinant plasmids; JT conceived and designed the experiments, analyzed the data, wrote a research proposal for funding and the manuscript.
This work was financially supported by the 2018–2019 fiscal year budget of the office of the National Research Council of Thailand to Thammasat University. PR received the Thammasat University Ph.D. scholarship.
Compliance with ethical standards
Consent for publication
All authors consent to publication.
Conflict of interest
All authors declare no personal or professional conflicts of interest, and no financial support from the companies that produce and/or distribute the drugs, devices, or materials described in this report.
Ethics approval and informed consent
This article does not contain any studies with human participants performed by any of the authors.
- Chang SC, Lin JY, Lo LY, Li ML, Shih SR (2004) Diverse apoptotic pathways in enterovirus 71-infected cells. J Neuro-Oncol 10:338–349Google Scholar