US-UK BBSRC-NIFA Collab: Evolution of the high pathogenicity phenotype in avian influenza virus

甲型流感病毒可以感染广泛的宿主，但其主要宿主是野生水禽。从这些宿主，病毒可以感染家禽，并引起家禽养殖业的特别关注，因为感染可能导致高死亡率。禽流感在家禽中的暴发表现出一系列疾病的严重性，从轻微症状到迅速死亡。特别令人关注的是高致病性禽流感(HPAI)的爆发。到目前为止，只有H5和H7禽流感亚型引起了HPAI暴发。HPAI可以直接从野鸟传入，但更常见的是与由前体低致病性禽流感(LPAI)引起的疾病严重性增加有关。高致病性表型的获得与HA中多碱裂解位点(PBCS)的引入有关。HA必须被切割才能被激活，而PBCS的存在意味着蛋白质可以被更广泛的宿主细胞蛋白酶激活，从而增加受病毒影响的组织，并导致全身传播和死亡。然而，在所有病例中，获得PBCS不足以增加病毒的致病性，一些具有PBCS的H5/H7病毒在禽类中表现出低致病性表型。因此，迫切需要了解影响获得HPAI表型的其他因素，以便更准确地评估与新出现的AIV感染相关的风险。我们认为，病毒离子通道蛋白M2和最近发现的一个称为M42的变体，通过一些机制在LPAI到HPAI的转变中发挥作用，本提议将探索这一假说。我们最近发现了一种名为M42的M2变异体，它在胞外结构域上与M2只有轻微的不同。这一微小的差异足以导致M42相对于M2的亚细胞定位转移到高尔基体。我们将检验这样的假设，即M42的亚细胞定位改变对于调节HA活性是重要的，并且M42的表达水平可能影响获得高致病性表型。M2已被提议形成用于人和家禽的通用疫苗的基础，该疫苗利用了M2的胞外区在病毒亚型之间是保守的这一事实。由于M42的胞外区与M2不同，在细胞培养研究中，M42的表达也被认为是流感病毒逃避M2抗体抑制的一种机制。到目前为止，还没有研究表明M42的表达是否可以为禽流感病毒提供一种克服这种疫苗接种策略的机制。在本建议中，我们将研究(I)禽流感病毒的M2/M42蛋白是否参与了从低致病性病毒到高致病性病毒的转变，以及(Ii)从M2到M42的改变是否对逃避疫苗反应具有重要意义。

Influenza A virus can infect a wide range of hosts, but its main reservoir is wild aquatic waterfowl. From these hosts, the virus can infect domestic birds and is of particular concern to the poultry industry, where infection can cause high mortality. This in turn can jeopardize the food supply chain, while outbreaks also have the potential to affect human health.Outbreaks of avian influenza (AI) in domestic poultry show a broad range of disease severity, from mild symptoms to rapid fatality. Of particular concern are outbreaks of High Pathogenicity Avian Influenza (HPAI). To date, only the H5 and H7 HA subtypes of AIV have caused HPAI outbreaks. HPAI can be introduced directly from wild birds, but is more commonly associated with the development of increased disease severity from a progenitor Low Pathogenicity Avian Influenza (LPAI). Acquisition of a high pathogenicity phenotype is associated with introduction of a polybasic cleavage site (PBCS) in HA. HA must be cleaved to be activated, and the presence of a PBCS means the protein can be activated by a broader range of host cell proteases, thus increasing the tissues affected by the virus, and leading to systemic spread and death. However acquisition of a PBCS is insufficient to increase the pathogenicity of the virus in all cases, and some H5/H7 viruses with a PBCS display a low pathogenicity phenotype in birds. Therefore there is an urgent need to understand other factors affecting acquisition of a HPAI phenotype in order to more accurately assess the risk associated with emerging AIV infections. We believe that the viral ion channel protein M2, and a recently identified variant of this called M42, play roles in the LPAI to HPAI transition through a number of mechanisms, and this proposal will explore this hypothesis. We recently identified an M2 variant called M42, which differs only slightly from M2 in its extracellular domain. This minor difference was enough to cause a shift in sub cellular localisation of M42 relative to M2, to the Golgi apparatus. We will test the hypothesis that the altered subcellular localisation of M42 is important for regulating HA activity, and M42 expression levels may affect the acquisition of a high pathogenicity phenotype. M2 has been proposed to form the basis of a universal vaccine for use in human and poultry which takes advantage of the fact that the extracellular domain of M2 is conserved amongst virus subtypes. Since the extracellular domain of M42 is different from M2, expression of M42 has also been identified as a mechanism by which influenza virus can escape from inhibition by antibodies targeted against M2 in cell culture studies. To date, no studies examining whether expression of M42 could provide a mechanism for AIV to overcome this vaccination strategy have been performed in birds.Overall, in this proposal we will investigate (i) if the M2/M42 proteins of avian influenza virus are involved in the change from low pathogenic to highly pathogenic virus and (ii) if the change from M2 to M42 is of importance to escape vaccine responses.