|dc.description.abstract||Influenza virus continues to pose serious medical and economic challenges to global public health as novel influenza viruses regularly emerge in human population. A universal influenza virus vaccine is not available yet and currently available influenza type- and subtype-specific vaccines need regular updating. Therefore, antiviral drugs are the first line of defence against a novel influenza virus. However, currently-approved anti-influenza drugs target viral components and influenza has mutated those components to acquire the drug resistance. Therefore, there is a need to develop alternative, effective, and long-lasting antiviral strategies to overcome the continuously emerging novel and drug-resistant influenza viruses in humans. One approach is to selectively target specific interactions of influenza virus with host pro-viral and anti-viral factors to inhibit virus replication. This strategy will less likely induce the viral resistance.
This PhD project was designed to determine the role of host histone deacetylase (HDAC) 1 and 2 in the infection of influenza A virus (IAV), the most significant influenza virus. The rationale of this project was based upon the recent discovery in our lab indicating that host histone deacetylases (HDACs) are potentially a family of novel anti-IAV factors. HDACs catalyse the deacetylation of a variety of cytoplasmic and nuclear proteins, consequently regulating diverse cellular processes. Mammalian HDACs have been divided into four classes. The HDAC1 and 2 belong to class I, and are the first and second discovered HDACs, respectively. Earlier, our lab demonstrated that HDAC6, a class II HDAC, possesses an anti-IAV property and IAV downregulates HDAC6 activity to potentially undermine its antiviral function. Based on these findings, we proposed the hypothesis that class I HDACs also have similar properties. HDAC1 is a prototypic member of class I and HDAC2 has about 86% amino acid sequence similarity with HDAC1. Hence, we investigated the role of HDAC1 and 2 in IAV infection using primarily the human lung epithelial cells and IAV PR/8/34(H1N1) and WSN/1933(H1N1) strains as a model.
We have found that IAV downregulates the HDAC1 expression (both at mRNA and polypeptide level) as well as its deacetylase activity, which, consistent with our hypothesis indicated an anti-IAV role of host HDAC1. Indeed, silencing of HDAC1 expression by RNA interference augmented the IAV infection by more than 6-fold, and conversely, the ectopic expression of HDAC1 from a plasmid decreased it by more than half. To efficiently replicate, IAV has evolved multiple strategies to circumvent the host innate antiviral response. The dysregulation of host HDACs could be part of that strategy as HDACs have been shown to be an important component of host innate response in a heterologous system. Consistent with this hypothesis, treatment of infected cells with trichostatin A (TSA), a widely-used HDAC inhibitor resulted in the downregulation of the phosphorylation of STAT1, a critical component of host innate antiviral response and expression of interferon-stimulated genes, IFITM3, ISG15, and viperin, which have been previously reported to have the anti-IAV function. Consequently, TSA treatment also resulted in the enhancement in IAV infection by more than 5-fold. Further, knockdown of HDAC1 expression resulted in decreased level of phosphorylated interferon regulatory factor 3, a key molecule in interferon signalling and subsequently the reduced expression of interferon α. Furthermore, the expression of viperin was also reduced or enhanced by about 58% or 55% in HDAC1-depleted or HDAC1-overexpressing cells, respectively.
Similarly, HDAC2 mRNA and polypeptide expression was also downregulated in IAV infected cells, albeit by a mechanism distinct to HDAC1 downregulation. Nevertheless, the knockdown of HDAC2 expression resulted in about 4 fold increase in IAV infection. In addition, there was a modest, but consistent decrease in the level of phosphorylated STAT1 in HDAC2-depleted cells and consequently, a decrease in viperin expression.
In summary, this PhD study has demonstrated an anti-IAV role of host HDAC1 and 2 and provided a significant insight into their antiviral mechanism. Evolutionary, HDAC1 and 2 are similar proteins and both were found to have the anti-IAV properties. However, HDAC1 and 2 seem to have a slightly distinct and independent interaction with IAV. Based upon the experimental evidence presented here, further mechanistic roles of these HDACs in IAV infection has been discussed and relevant future research directions have been outlined. In conclusion, both HDAC1 and HDAC2 provide a cellular refractory state to IAV infection by regulating the host innate immune response. The data presented here will contribute to further molecular understanding of the IAV-HDACs interactions.||