Prion Protein as a Modulator of Influenza Sakaguchi and Chida

Worldwide spread of influenza A virus (IAV) strains, which are resistant to currently available antiinfluenza agents such as viral neuraminidase inhibitors, has encouraged identification of new target molecules for anti-influenza agents. Reactive oxygen species (ROS) causing oxidative stress play a pivotal role in the pathogenesis of lung injuries induced by infection with IAVs, therefore suggesting that anti-oxidative therapeutics targeting cellular molecules could be beneficial against IAV infection without inducing drug-resistant IAV strains. We recently found that the normal cellular prion protein, PrPC, whose conformational conversion into the amyloidogenic isoform, PrPSc, in the brain is a key pathogenic event in prion diseases, is expressed by lung epithelial cells and exerts a protective role against IAV infection in mice by reducing ROS in infected lungs. The Cu content and activity of antioxidative enzyme Cu/Zn-superoxide dismutase, or SOD1, were lower in the lungs of PrPC-knockout mice, suggesting that the anti-oxidative activity of PrPC is probably attributable to its function of activating SOD1 through regulating Cu content in lungs. Here, we introduce PrPC as a novel modulator of influenza and its potential implication for anti-oxidative therapies for IAV infection. We also introduce other candidate targets reported for antioxidative anti-influenza therapies. Introduction Influenza A viruses (IAVs) are enveloped, negative sense, single-stranded RNA viruses, causing seasonal epidemics of influenza affecting about 20% of the world population annually, with 250,000-500,000 deaths each year (Fiore et al., 2008). The young and elderly and those with underlying chronic diseases in lung or cardiovascular systems are particularly vulnerable to IAV infection (Fiore et al., 2008). There have been so far four pandemics of influenza in the past; the 1918-1919 Spanish flu pandemic caused by infection with IAV subtype H1N1, the 1957-1958 Asian flu pandemic by H2N2, the 1968-1970 Hong Kong flu pandemic by H3N2, and the 2009-2010 swine flu pandemic by H1N1, causing greater than 50 million deaths worldwide (Nickol and Kindrachuk, 2019). Potential risks of emergence of new IAV subtypes and their worldwide spread leading to new flu pandemics are still high. In addition, many sporadic infections with highly pathogenic avian influenza virus (HPAIV) subtype H5N1 to humans, causing high morbidity and mortality in infected people, have been reported (Nunez and Ross, 2019). As of July 2018, a total of 860 cases of H5N1 infection in humans have been confirmed and 454 deaths have been reported (Nunez and Ross, 2019). There have been only a few cases of humanto-human transmission of HPAIVs (Nunez and Ross, 2019), suggesting a very low likelihood of pandemic threat of H5N1 infection in human populations. However, there are potential risks that HPAIVs might change their pathogenic properties in humans and produce new types of progeny viruses, which are more susceptible to humans and more easily spread among human populations through human-to-human infection, eventually causing new flu worldwide in the world. 21 Curr. Issues Mol. Biol. (2020) 37: 21-32. caister.com/cimb Prion Protein is a Novel Modulator of Influenza: Potential Implications for Anti-Influenza Therapeutics Prion Protein as a Modulator of Influenza Sakaguchi and Chida Vaccines against the viral surface glycoprotein hemagglutinin (HA) are currently available as a prophylactic measure against IAV infection (Yamayoshi and Kawaoka, 2019). However, they are only effective if they target the current circulating viruses. Furthermore, they also cannot reduce the ongoing threat of new pandemic strains since production of the vaccines takes a long time. Inhibitors of the viral neuraminidase (NA), which is essential for release of IAV progenies from infected cells, are the currently available major antiviral agents (Robson et al., 2019). The efficacy of the agents has been clearly demonstrated. However, the agents need to be used within 48 hours of onset and are most effective within 24 hours (Robson et al., 2019). In addition, certain strains of IAVs have already acquired resistance to these anti-influenza agents and have been spreading in human populations worldwide (Robson et al., 2019). The anti-influenza agents target the viral molecules, thereby stimulating the emergence of drug-resistant IAVs, which carry mutations in the genes encoding those molecules. Therefore, identification of new target molecules for anti-influenza agents is urgently awaited. Many cellular molecules play a crucial role in the pathogenesis of IAV infection (Marques et al., 2019), suggesting that these cellular molecules could be plausible targets for new anti-influenza agents. Lines of evidence indicate that oxidative stress plays a pivotal role in the pathogenesis of IAV infection (Akaike et al., 1990; Oda et al., 1989; Vlahos and Selemidis, 2014). We recently identified that the normal cellular prion protein, PrPC, could have a protective role against lethal infection with IAVs in mice by functioning as an anti-oxidative molecule by regulating an anti-oxidative enzyme in lungs (Chida et al., 2018). In this review, we introduce PrPC as a novel potential candidate target for anti-oxidative therapeutics against IAV infection, as well as other candidate targets reported for antioxidative anti-influenza therapeutics. Brief overview of IAV infection IAV infection begins with attachment of the virus to the surface of the airway or lung epithelial cells and alveolar macrophages through specific binding of the viral surface protein HA to either α2,3or α2,6linked sialic acids expressed on the cell surface (Chutinimitkul et al., 2010; van Riel et al., 2010). Seasonal and pandemic IAV strains use α2,6-linked sialic acids for binding to the host cells while HPAIV strains use α2,3-linked sialic acids (Shinya et al., 2006; van Riel et al., 2010). α2,6-linked sialic acids are prominently expressed in the trachea, whereas α2,3-linked sialic acids are expressed by alveolar type II (ATII) epithelial cells (Shinya et al., 2006; van Riel et al., 2010). This specific expression of α2,3linked sialic acids in ATII cells, which is located in the respiratory tract terminus, alveolus, have been suggested to explain why HPAIV strains have difficulty infecting humans. Binding of IAVs to the cells triggers endocytosis of the virus into them. The acidic environment of the endosomes causes fusion of the virus with the endosomal membrane, resulting in release of the viral genome into the cells eventually leading to propagation of the viral progenies (Pinto and Lamb, 2006). The M2 viral protein is activated in the endosomes and forms an ion channel on the viral membrane, allowing entrance of protons into the viral core to dissociate the ribonucleoprotein complex (Pinto and Lamb, 2006). The newly replicated viral progenies are still bound to the host cells through linkage of HA of the progenies to sialic acids on the cell surface (Rossman and Lamb, 2011). The viral protein NA cleaves the link, allowing release of the progeny viruses from the host cells and spread of IAV infection in the lungs (Rossman and Lamb, 2011). IAV infection can be cleared by the host immune systems. The innate immune system detects IAV infection through recognition of the virus by host cells via interaction of pathogen-associated molecular patterns, which are present in the virus or generated during infection, with cellular pattern recognition receptors such as the Toll-like receptors (TLRs), retinoic acid-inducible gene I (RIG-I), and NOD-, LRRand pyrin domain-containing 3 (NLRP3) (Iwasaki and Pillai, 2014). TLR3 and TLR7 recognizes dsRNA and ssRNA in endosomes, respectively, and RIG-I detects 5'-triphosphate viral ssRNA that is generated in the cytoplasm after IAV infection (Iwasaki and Pillai, 2014). These interactions between viral and cellular molecules trigger activation of innate immunity against IAV infection, leading to production of pro-inflammatory cytokines, recruiting immune cells into the infection sites, activating adaptive immune responses against IAV infection, eventually eliminating IAV infection (Iwasaki and Pillai, 2014). NLRP3 is expressed in monocy tes , dend r i t i c ce l l s , neu t roph i l s , macrophages, and epithelial cells and forms the multiprotein complexes called inflammasomes together with the ASC protein and pro-caspase 1 (Guarda et al., 2011). IAV infection triggers f o rma t i on and ac t i va t i on o f t he NLRP3 i n f l ammasomes , resu l t i ng i n re lease o f interleukin-1β (IL-1β) and IL-18 and thereby