Identification of receptors for H5N1 virus on human nerve cells using protromics-based approaches

Abstract

In this study, we investigated neuroinfectious capacity of H5N1 (A/Thailand/NK165/2005) isolated from plasma of infected individual during the third wave of Thailand outbreaks using human neuroblastoma SH-SY5Y cells. This was due to lack of information regarding neuroinfectivity of this variant. Results demonstrated that H5N1/NK165 induced servere CPEs in these neuronal cells. H5N1-specific hemagglutinin was found in the cytoplasm of the infected cells as early as 12 h post-infection. By 24 h post-infection, all cells in the cultures were infected. These findings coincided with time course of CPEs and the presence of virus progeny in the culture medium of infected cells. Virus titer increased exponentially overtime. After 24 h post-infection, viability of the infected cell began to decline. By 72 h, only cellular debris remained in the infected cultures. Moreover, with this isolate we identified host cell proteins that might be involved in H5N1 entry to nerve cells using 1D-VOPBA coupled with LC-MS/MS analysis. RACK1 and prohibitin were identified as potential receptor for H5N1. Expression of RACK1 and prohibitin was confirmed by western blot analysis. There was a significant decrease in Expression of RACK1 and prohibitin preteins compared with mock-infected cells, suggesting that both RACK1 and prohibitin might be involved in the initial step of H5N1 binding to the neuronal cell membrane and/or internalization. Further studies are needed to explore whether both RACK1 and prohibitin are actual receptors for H5N1. This will ultimatel lead to improved therapeutics and provide an insight into neuronal mechanism of H5N1 infection.