Caister Academic Press


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A review of scientific research into Influenza.


Adapted from Qinghua Wang and Yizhi Jane Tao in Influenza: Molecular Virology
In the last 100 years there have been three major influenza pandemics: Spanish Flu in 1918, Asian Flu in 1957 and Hong Kong Flu in 1968. These claimed the lives of approximately 50 million, 2 million and 1 million people respectively. Added to this is the annual death toll of 250,000 to 500,000 people worldwide with a further 3 to 4 million people suffering severe illness. These statistics make influenza an extremely important pathogen. In 1997 the alarming emergence of a new, highly pathogenic subtype, H5N1, which has a 50% mortality rate, provided a major impetus for renewed influenza research. However the battle against influenza is going to be difficult. Recently another subtype, H1N1, has emerged. This subtype causes a relatively mild infection in humans, however is highly transmittable between people and a new influenza pandemic was declared by the World Health Organization. If this virus were to acquire some of the lethal capabilities of H5N1, then the ensuing pandemic could be devastating.

The 1918 Pandemic Influenza Virus

Adapted from Natalie Pica, Terrence M. Tumpey, Adolfo García-Sastre and Peter Palese in Influenza: Molecular Virology
The pandemic influenza virus of 1918 was extremely virulent and caused significant morbidity and mortality to millions of people worldwide. The extinct virus caused severe pathology in both the upper and lower respiratory tract, resulting in fatal respiratory complications and bacterial pneumonia. The pathology associated with 1918 influenza virus infections is thought to be the result of the exposure of an immunologically naive host population to an unusually virulent virus. Using reverse genetics, the 1918 pandemic virus has been studied in an attempt to determine which viral genes contribute to the increased virulence. Studies to date point to the role of the hemagglutinin, neuraminidase, and the polymerase basic protein 1 genes as the virulence genes responsible for the high pathogenicity seen with the 1918 influenza virus.

A Diagnostic Influenza Microarray

Adapted from Erica D. Dawson and Kathy L. Rowlen in Influenza: Molecular Virology
Rapid and accurate diagnostic methods for typing and subtyping influenza viruses are needed for improved worldwide surveillance. Although molecular-based diagnostic methods are becoming more widespread in influenza diagnosis, they generally involve amplification of the hemagglutinin (HA) and/or neuraminidase (NA) gene segments for subtyping. Lab-on-a-chip microarrays have been developed (MChip) that allow for the identification and subtyping of influenza A viruses in approximately seven hours. MChip is unique in that it is based solely on the matrix (M) gene segment which has enough genetic diversity for subtype analysis but sufficient genetic stability to circumvent the need for continual redesign of primers and microarray probes. An overview is presented here of several MChip studies undertaken over the last several years, highlighting the ability of MChip to identify and subtype influenza A/H1N1, H3N2, and H5N1 viruses.

Influenza Vaccines

Adapted from Hao Zhou, Ramdas S. Pophale and Michael W. Deem in Influenza: Molecular Virology
A new parameter has been defined to quantify the antigenic distance between two H3N2 influenza strains and this parameter has been used to measure antigenic distance between circulating H3N2 strains and the closest vaccine component of the influenza vaccine. For the data between 1971 and 2004, the measure of antigenic distance correlates better with efficacy in humans of the H3N2 influenza A annual vaccine than do current state of the art measures of antigenic distance such as phylogenetic sequence analysis or ferret antisera inhibition assays. This measure of antigenic distance could be used to guide the design of the annual flu vaccine. The measure of antigenic distance can be combined with a multiple-strain avian influenza transmission model to study the threat of simultaneous introduction of multiple avian influenza strains. For H3N2 influenza, the model has been validated against observed viral fixation rates and epidemic progression rates from the World Health Organization FluNet - Global Influenza Surveillance Network. A multiple-component avian influenza vaccine is helpful to control a simultaneous multiple introduction of bird-flu strains. A multiple vaccine may offer improvements.

The Influenza NS1 Protein

Adapted from Zachary A. Bornholdt, Berenice Carrillo and B.V. Venkataram Prasad in Influenza: Molecular Virology
The non-structural protein 1 (NS1) of influenza virus is a potent antagonist of the cellular antiviral interferon (IFN) response. It is a multifunctional protein with two domains, a dsRNA binding domain (RBD) and an effector domain (ED) which interacts with various cellular proteins. Although, initially sequestration of dsRNA was considered the primary mechanism for countering IFN, subsequent studies have shown that the interactions of ED with various cellular proteins are likely involved. NS1 is shown to be a virulence determinant, especially in the highly pathogenic H5N1 viruses that are currently a threat for another influenza pandemic. Among various influenza virus strains, NS1 is relatively well conserved with major differences occurring in the linker region and the C-terminus, where several NS1 proteins contain truncations. How these differences contribute to virulence remains unknown but these differences seem to have an effect on NS1 function that may be strain specific. Substantial progress has been made toward understanding of the structural aspects of this two-domain protein.

Adapted from Chen Zhao, Rei-Lin Kuo and Robert M. Krug in Influenza: Molecular Virology
The NS1 protein of influenza A viruses is a small, multi-functional dimeric protein that participates in both protein-RNA and protein-protein interactions. It is comprised of two functional domains: N-terminal RNA-binding domain and C-terminal effector domain. A major role of the NS1 protein is to counter host cell antiviral responses. The RNA-binding domain binds double-stranded (ds) RNA, thereby inhibiting the dsRNA activation of the antiviral oligo A synthetase/RNase L pathway that is induced by interferon-α/β (IFN-α/β). A region of the effector domain binds the protein kinase PKR, thereby preventing its activation that would otherwise lead to the shutdown of both viral and host protein synthesis. Another region of the effector domain binds the 30 kDa subunit of the cleavage and polyadenylation specificity factor (CPSF30), a cellular protein required for the 3' end processing of all cellular pre-mRNAs. As a consequence, the substantial amount of IFN-β pre-mRNA that is synthesized in virus-infected cells is not processed to form mature IFN-β mRNA, thereby suppressing the IFN response. The NS1 protein also has other functions that are not directly involved in countering host antiviral responses. The effector domain of the NS1 protein binds the P85β regulatory subunit of phosphoinositide 3-kinase (PI3K), resulting in the activation of PI3K and the Akt kinase, which in turn inhibits apoptosis. The C-termini of pathogenic influenza A viruses have a PDZ-binding motif that has been implicated in pathogenicity. The NS1 protein also interacts with the cellular nuclear export protein (TAP), and may have a role in the nuclear export of viral mRNAs. Finally, the NS1 protein functionally interacts with the viral polymerase complex in infected cells and likely has a role in the regulation of viral RNA synthesis.

Influenza Virus Nucleoprotein

Adapted from Yizhi Jane Tao and Qiaozhen Ye in Influenza: Molecular Virology
The (-) RNA genome of the influenza A virus, eight segments in total, is encapsidated in the form of ribonucleoprotein (RNP) complexes. The nucleoprotein (NP), the major protein component of RNPs, binds along the entire length of each genomic RNA segment at a 24-nt interval, forming the double-helical RNP structures found in mature viruses. The viral polymerase, consisting of PA, PB1, and PB2 subunits, binds to the two RNA termini of the RNP. As one of the most abundant proteins made in infected cells, influenza virus NP has essential roles in many important viral processes, including intracellular trafficking of the viral genome, viral RNA replication, viral genome packaging, and virus assembly. The recently determined crystal structures of two NP trimers show an overall fold and an external RNA binding mode that are different from rhabdovirus NP, as confirmed by a new cryo-EM reconstruction of a mini-RNP. Site-directed mutagenesis and RNA binding assays have confirmed that a positively charged groove plays an important role in NP:RNA binding.

Influenza Virus Hemagglutinin Glycoproteins

Adapted from David A. Steinhauer in Influenza: Molecular Virology
The influenza A virus hemagglutinin glycoprotein (HA) is the principle mediator of viral entry into host cells. It is responsible for attachment of virions to sialic acid-containing receptors on the host cell surface, and for inducing membrane fusion between viral envelopes and cellular endosomal membranes following endocytosis. HA serves a classic example of a type I membrane glycoprotein, with a cleaved N-terminal signal sequence, a membrane anchor domain near the C-terminus, and post-translational modifications resulting from the addition of N-linked oligosaccharide side chains to the ectodomain, and acylation of cysteine residues in the cytoplasmic tail region. HA spikes on the viral surface are also the major target for neutralizing antibodies, and as such, the antigenic properties of the HA are of fundamental significance for the design of influenza vaccines. The depth of knowledge relating to high resolution atomic structures of HA in various forms have made it a prototype for the investigation of viral glycoproteins in general.

Influenza M2 Channel

Adapted from James J. Chou and Jason R. Schnell in Influenza: Molecular Virology
Viral ion channels have minimalist architecture. Despite their relatively simple structure, viral channels can achieve highly specific gating and selection of ions, and the particular mechanisms appear to be different from those of prokaryotes and eukaryotes. The unique structural and functional properties of viral channels make them ideal targets for antiviral therapy because the molecules that inhibit viral ion channels may not interact with human ion channels. The M2 proton channel of influenza A virus is a model viral ion channel. This small channel, whose pore is formed by four equivalent transmembrane helices, is the target of two widely used anti-influenza A drugs, amantadine and rimantadine, both belonging to the adamantane class of compounds. However, resistance of influenza A to adamantane is now widespread. Naturally-occurring resistants mutants have been observed in as many as six different positions in the transmembrane segment of M2. How could there be so many different resistance-conferring mutations along a transmembrane helix of 25 amino acids? The recently-determined high-resolution structures of M2 in complex with adamantane allow us to begin answering this question.

Influenza Polymerase

Adapted from Mark Bartlam, Zhiyong Lou, Yingfang Liu and Zihe Rao in Influenza: Molecular Virology
The influenza virus RNA-dependent RNA polymerase is a heterotrimeric complex (PA, PB1 and PB2) with multiple enzymatic activities for catalyzing viral RNA transcription and replication. Its critical roles in the influenza virus life cycle and high sequence conservation suggest it should be a major target for therapeutic intervention. Functional studies and drug discovery targeting the influenza polymerase have been hampered by the lack of three-dimensional structural information. There has been recent progress in study of the structure and function of the influenza polymerase, and there may be prospects for the development of anti-influenza therapeutics.

Further reading

Further reading