Protein Disulfide Isomerase A3 Regulates Influenza Neuraminidase Activity and Influenza Burden in the Lung.

Emily M Nakada,Zoe F Mark,Danielle Antos,Sierra R Bruno,Dhemerson Souza De Lima,Nicolas Chamberlain,John F Alcorn,Mona Ruban,Vikas Anathy,Ravishankar Chandrasekaran,Amit Kumar

doi:10.3390/ijms23031078

Abstract

Influenza (IAV) neuraminidase (NA) is a glycoprotein required for the viral exit from the cell. NA requires disulfide bonds for proper function. We have recently demonstrated that protein disulfide isomerase (PDI)A3 is required for oxidative folding of IAV hemagglutinin (HA), and viral propagation. However, it not known whether PDIs are required for NA maturation or if these interactions represent a putative target for the treatment of influenza infection. We sought to determine whether PDIA3 is required for disulfide bonds of NA, its activity, and propagation of the virus. Requirement of disulfides for NA oligomerization and activity were determined using biotin switch and redox assays in WT and PDIA3−/− in A549 cells. A PDI specific inhibitor (LOC14) was utilized to determine the requirement of PDIs in NA activity, IAV burden, and inflammatory response in A549 and primary mouse tracheal epithelial cells. Mice were treated with the inhibitor LOC14 and subsequently examined for IAV burden, NA activity, cytokine, and immune response. IAV-NA interacts with PDIA3 and this interaction is required for NA activity. PDIA3 ablation or inhibition decreased NA activity, viral burden, and inflammatory response in lung epithelial cells. LOC14 treatment significantly attenuated the influenza-induced inflammatory response in mice including the overall viral burden. These results provide evidence for PDIA3 inhibition suppressing NA activity, potentially providing a novel platform for host-targeted antiviral therapies.

Highlights

The influenza A virus (IAV) causes severe respiratory illness and worldwide impact [1].While vaccination and therapeutics are available, they are often rendered ineffective due to the accumulation of mutations in the viral genome [1]
Disulfide Bonds Are Required for NA Activity, and PDIA3 Interacts with IAV NA
The production of IFNβ was decreased in PDIA3−/− compared to wild type (WT) cells (Figure 1K). These results indicated that PDIA3 and NA interact, and disulfide bond formation in NA depends on PDIA3

Summary

Introduction

The influenza A virus (IAV) causes severe respiratory illness and worldwide impact [1]. While vaccination and therapeutics are available, they are often rendered ineffective due to the accumulation of mutations in the viral genome [1]. One potential strategy to circumvent this type of mutational resistance is targeting the host proteins or post-translational processes utilized by the virus during propagation. IAV and host protein disulfide isomerases (PDIs), involved in disulfide bond formation, is emerging, and viruses often utilize redox-active host chaperone proteins to assist in their protein folding [2–7]. PDIs are a major disulfide catalyzing family of enzymes in mammalian cells [8]. PDIA3, is known to be required for efficient folding of IAV hemagglutinin (HA)

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Results

Conclusion