Influenza virus
1.1.1 Viruses: general information
The word virus is from the Latin noun virus, meaning toxin or poison. Viral diseases such as rabies, yellow fever, and smallpox have affected humans for many centuries. There is evidence of polio in the ancient Egyptian empire, though the cause of these diseases was unknown at the time.
Viruses are classified as either a DNA or RNA virus according to the nucleic acid type of their genetic material. The RNA viruses are divided into three groups: Group I - viruses possessing double-stranded RNA genomes, e.g. rotavirus; Group II - viruses possessing positive-sense single-stranded (dsRNA) RNA genomes including for example Hepatitis A virus, enteroviruses, rhinoviruses, poliovirus, foot-and-mouth virus and Group III - viruses possessing negative-sense single-stranded RNA genomes, including for example, the deadly Ebola and Marburg viruses, the influenza virus, measles, mumps and rabies viruses. Some negative sense RNA viruses contain also positive sense RNA and are referred to as ambisense viruses (e.g. some bunyaviruses)1.
Viruses lack the cellular machinery for self-reproduction. The viral genome codes for the proteins that constitute the protective outer shell (capsid) as well as for those proteins required for viral reproduction that are not provided by the host cell. The capsid consists of monomeric subunits of protein and serves to protect the virus's genetic material. The capsid or the membrane (in the case of membrane viruses) target cells suitable for infection, and initiate the infection by "attaching" to the target cell, followed by uptake of the virus and release of the DNA or RNA into the cytoplasm. After entering the cell, the virus's genetic material begins the process of parasitizing the cell to produce new virions1.
1.1.2 Influenza
The highly contagious, acute respiratory illness known as influenza, appears to have afflicted human kind since ancient times. The word Influenza come from the Italian form of Latin influentia, meaning “inflow”, originally used because epidemics were thought to be due to an influx, or influence, of astrological or other occult sources.
The Influenza virus is a negative sense (group III) RNA virus of the family Orthomyxoviridae. There are three types of influenza viruses: Influenza A virus, Influenza B virus, and Influenza C virus. Influenza A and C infect multiple species, while Influenza B infects almost exclusively humans. My research has focused on Influenza A.
The major reservoir for Influenza A virus is in birds, but it may also infect several species of mammals. Unusual for a virus, the Influenza A virus genome is not a single segment of nucleic acid; instead, it contains eight segments of negative-sense RNA (13.5 kilo bases total), which encode for 11 proteins2.
Although occasionally occurring as sporadic infections, influenza is more commonly and dramatically seen as local outbreaks or widespread epidemics: these occur in most parts of the world. Epidemics can arise at any time but are usually concentrated to months of relatively high humidity. They occur suddenly and unexpectedly, often with little or no warning, and the number of people infected can vary from a few hundred to hundreds of thousands. History records nine occasions since 1700 AD when influenza has caused pandemics, and at these times, millions were infected. In many cases, epidemics have been short-lived, lasting days or weeks; however, those occurring in large groupings can continue in successive waves for months and even years. The large number of persons infected during an outbreak of influenza, together with our proven inability to prevent or to contain these outbreaks, is the reason that so much study and research has focused on this disease3.
Epidemic Influenza is a short-lived but relatively severe respiratory infection in healthy adults. The large number of patients involved in an epidemic can include a significant number of deaths. Epidemics can be disruptive to industry, with loss of productivity, and to services such as medical, postal, power, police and education. It can cause depressed immunity to other infectious diseases; it can cause depression and behavioral complications, which may continue for months after the acute phase of illness has passed. Infection is a life threatening condition to the elderly, babies and to patients with predisposing heart, chest, or metabolic disease1.
Epidemics can be traced anecdotally from the fifth century BC, the Hundred Years War and the court of Mary Tudor, to the more exact records of the last 100 years. Nevertheless, knowledge gained during this latter period, especially the past 70 years when the virus could be isolated and studied in the laboratory, has done little to prevent epidemics or help treat patients. It is not surprising therefore, that influenza has been referred to as the ‘last plague’. Since the first isolation of an influenza virus in ferrets in 1933, research into the nature and control of the disease has led to the setting up of many research laboratories devoted to the study of influenza viruses, and to the commissioning of an international network of communicating laboratories by the World Health Organization (WHO) to monitor the antigenic changes of the infecting viruses and the incidence and spread of infection3. As a result of these activities, a large volume of information has become available concerning influenza viruses. Whether application of this knowledge can be used to diminish the impact of influenza on individuals or to prevent the epidemics and pandemics which will inevitably occur in the future remains to be seen3.
Influenza viruses, while examined under an electron microscope show spherical particles, with a diameter of 80–120 nm. The virus consists of ssRNA of negative polarity 1, segmented into eight fragments. The RNA is closely associated with the nucleoprotein (NP) to form the ribonuclear protein (RNP), a helical structure termed the nucleocapsid. Surrounding the nucleocapsid is a second protein referred to as the matrix or membrane protein (M1); this protein is the major protein in the virus particle (approximately 3000 molecules in each virus particle), contributing 35–45% of the particle mass. There is a second M protein, termed M2, which is coded by the same gene segment as the M1 protein. M2 is important in viral replication. External to the M1 protein the virion particles are surrounded by the viral membrane; this is a lipid bilayer, which constitutes approximately 20–24% of the virus particle mass.
Two virus glycoproteins are inserted into the membrane; these are rod-shaped structures radiating out from the virus particles to give the spiky appearance of the surface. The first of these glycoproteins is the haemagglutinin (HA), which is composed of two separate molecules, termed HA1 and HA2, joined together by a disulphide bond 4. The function of the HA is the attachment of the virus to receptors on the surface of host cells during the initial stages of virus infection and for the fusion of the virus membranes with that of the endocytic vesicles. The second glycoprotein radiating from the surface of the virus particle is the neuraminidase (NA). The function of the NA includes the removal of sialic acid (the virus's receptor) residues from cell surfaces to promote virus absorption, and thereby mediate release of newly formed virus particles from the cell surface 5.
Influenza virus particles also contain an RNA dependent RNA polymerase (RDRP) complex; this complex consists of three proteins (P-proteins), termed polymerase basic protein 1 (PB1), polymerase basic protein 2 (PB2) and polymerase acidic protein (PA). These P-proteins have a complex role in virus-specific RNA synthesis during viral replication. Finally, the virus RNA codes for two non-structural proteins, termed NS1 and NS2: NS1 is synthesized early after infection, while NS2, now also termed nuclear export protein (NEP), appears late and is found in virus particles 6. The function of these molecules remains an important subject of current research and it has been found that mutant viruses with defects in the RNA fragment coding for these proteins are unable to synthesize sufficient viral RNA or M1 proteins. However, research to date implicates NS1 as a down-regulator of interferon production and NS2 as a facilitator of nuclear export of newly synthesized copies of the influenza virus genomic segments.
1.1.5 Influenza Life cycle
1.1.5.1 Viral Penetration
The influenza virus binds specifically to the cell surface of certain cells through sialic acid-containing glycoproteins, which undergo interaction with the virus HA. Some 28 species of sialic acid receptors, together with sugar chains, are known 7, and the specificity is important, since avian influenza viruses bind to some receptors while human influenza viruses bind to others; thus, receptor characteristics are important in determining virus-host range 8. However, this is not absolute, since pathogenicity is affected by genetic determinants and some viruses can cross species barriers via attaching to several receptors 9.
The binding function of the HA was shown in studies using specific antibodies to the HA, which prevents virus adsorption. Viral adsorption provokes receptor-mediated endocytosis of the viral particles via clathrin-coated pits, which can be seen by electron microscopy to appear approximately 20 min after infection. This in turn is followed by fusion of the viral envelope to the endosomal membranes, a procedure which requires the acidic environment in the endosomes (pH5.0–6.0) and the cleavage of the HA into HA1 and HA2. The low pH also frees the M1 protein from the RNP, thus allowing the RDRP complex to migrate to the cell nucleus. The acidic environment within the virus particle that allows this is achieved via the M2 protein located in the virus membrane. M2 acts as a channel to allow H+ ions into the virion from the endocytic environment, which is also facilitated by lysosomal proteases and lipase enzymes and mediated by the NP 10.
The events of virus replication in the cell nucleus are complex and remain to be fully understood 1. Our information to date suggests that transcription of the messenger RNA (mRNA) from all RNA fragments takes place immediately after entry into the cell nucleus. First, the polymerase complex binds to a methylated cap structure present on heterologous, cellular mRNA; this cap recognition is a function of the PB2 protein. Second, a nucleotide sequence of 10–30 bases is cleaved from the cap structure by endonuclease activity, termed ‘cap snatching’. This structure forms the starter molecule for viral mRNA synthesis: it has been suggested that this cleavage is one of the functions of PB1 or PB2 proteins 11. The cap sequence incorporates a G complementary to the penultimate C of the vRNA to complete the initiation step, and this is a function of the PB1 protein. Finally, elongation takes place in the normal manner to produce the complete mRNAs; this step requires binding of the RDRP to NP. Elongation is thought to be controlled by the PA protein. It is evident that interactions occur between PB1 and PB2, and between PB1 and PA but none has been demonstrated between PB2 and PA to date.
The mRNAs leave the cell nucleus, bind to ribosomes and translation of viral proteins is initiated. The synthesis of complement RNA (cRNA), and subsequently viral RNA (vRNA), occurs after the peak production of mRNA and protein synthesis. The synthesis of cRNA and vRNA requires the NP to switch to a replicase mode. In contrast to the production of mRNA, cRNA and vRNA are full-length copies of the RNA; it is neither capped nor polyadenylated and does not require a primer12.
1.1.5.2 Viral Assembly and Release
Assembly of new virions within the infected cells begins with the binding of NP to newly synthesized vRNAs to form the RNP 13. RNPs exit the nucleus in association with M1 and Nuclear Export Protein (NEP), and at the cell membrane they are enclosed by an envelope which contains HA, NA and M2 proteins; it is proposed that the M1 protein is the major driving force in this process 14. Virus escapes by budding from lipid rafts in the plasma membrane: the process of virus release is detectable 5–6 h after infection. The mechanism for viral release from the host cell is not completely understood, but an important element in this process is believed to be the NA, which prevents limitation of virus release through aggregation, by the removal of sialic acid residues from the viral envelope and at the cell membrane12.
Refrences
1 RM, Lamb RA and Krug, Field’s Virology. (Lippincott-Raven, Philadelphia, PA, 1996).
2 Neumann, G., Watanabe, T., and Kawaoka, Y., Plasmid-driven formation of influenza virus-like particles. J Virol 74 (1), 547 (2000).
3 Kitler, M. E., Gavinio, P., and Lavanchy, D., Influenza and the work of the World Health Organization. Vaccine 20 Suppl 2, S5 (2002).
4 Skehel JJ, Daniels RS, Douglas AR, The Molecular Biology and Epidemiology of Influenza. (Academic Press, London, 1984).
5 P, Colman, Textbook of Virology. (Blackwell, London, 1998).
6 Paragas, J. et al., Influenza B and C virus NEP (NS2) proteins possess nuclear export activities. J Virol 75 (16), 7375 (2001).
7 Suzuki Y, Ito T, Suzuki T, Options for the Control of Influenza. (Excerpta Medica, Amsterdam, 2001).
8 Matrosovich, M. and Klenk, H. D., Natural and synthetic sialic acid-containing inhibitors of influenza virus receptor binding. Rev Med Virol 13 (2), 85 (2003).
9 Stray, S. J., Cummings, R. D., and Air, G. M., Influenza virus infection of desialylated cells. Glycobiology 10 (7), 649 (2000).
10 Wang, P., Palese, P., and O'Neill, R. E., The NPI-1/NPI-3 (karyopherin alpha) binding site on the influenza a virus nucleoprotein NP is a nonconventional nuclear localization signal. J Virol 71 (3), 1850 (1997).
11 Blok, V. et al., Inhibition of the influenza virus RNA-dependent RNA polymerase by antisera directed against the carboxy-terminal region of the PB2 subunit. J Gen Virol 77 ( Pt 5), 1025 (1996).
12 A. J. Zuckerman, J. E. Banatvala, J. R. Pattison, P. D. Griffiths and B. D. Schoub, Principles and Practice of Clinical Virology (John Wiley & Sons Ltd, 2004).
13 Elton, D., Medcalf, E., Bishop, K., and Digard, P., Oligomerization of the influenza virus nucleoprotein: identification of positive and negative sequence elements. Virology 260 (1), 190 (1999).
14 Gomez-Puertas, P. et al., Influenza virus matrix protein is the major driving force in virus budding. J Virol 74 (24), 11538 (2000).
Comments
Write New Comment ▼
Write New Comment
Sorry! This knol's owner(s) have blocked you from editing, making suggestions, or commenting here.