Ns2 122

Source: Adapted in part from Fields et al. (1996, Table 2, p. 1355).

Source: Adapted in part from Fields et al. (1996, Table 2, p. 1355).

it also activates the RNA polymerase of the virion through the activity of M2. M2 is the target of the drug amanta-dine, one of the relatively few drugs that are effective against a viral disease. Amantadine binds M2 and prevents it from acting as an ion channel, which prevents the activation of the polymerase. When taken early during infection, amantidine ameliorates the symptoms of influenza.

Influenza Glycoproteins

Comparison of the glycoproteins of influenza A virus and the paramyxovirus SV-5 is of interest. In both influenza A virus and SV-5, one of the glycoproteins is type 1 (N terminus out) and one is type 2 (C terminus out). In both cases, the type 1 glycoprotein is produced as a precursor that must be cleaved to activate the fusion activity required for entry into cells. The type 1 glycopro-tein of influenza A has fusion and receptor-binding (hemagglutinating) activities and is called the hemagglu-tinin or HA. The precursor is called HA0 and the cleaved products are called HA1 and HA2 (which remain cova-lently linked by a disulfide bond after cleavage of the peptide bond). The SV-5 type 1 glycoprotein has only fusion activity and is called F. As described above, it is produced as a precursor F0 which is cleaved to F1 and F2. The receptor bound by both influenza A virus and by SV-5 for entry into cells is sialic acid. The type 2 glycoprotein of influenza has neuraminidase activity and is called the neuraminidase or NA. It removes sialic acid from glyco-proteins for the same reasons as described for the paramyxoviruses that use sialic acid as a receptor. The type 2 glycoprotein of SV-5 has both neuraminidase activity and receptor-binding (hemagglutinating) activities and is called HN.

Influenza HA is present as a trimer on the surface of the virus (as is F of SV-5). The trimeric spike has a long stalk and a head containing the sialic acid binding sites. As described in Chapter 1, exposure to acid pH in endosomes produces a dramatic rearrangement of the spike in which the fusion peptide, which forms the N terminus of HA2, is moved over a distance of more than 10 nm to the tip of the spike (Fig. 1.5). Here it inserts into the target membrane and somehow promotes fusion of the viral membrane with the target membrane. NA is present as a tetramer (as is HN of SV-5), and forms a spike that is distinguishable in the electron microscope from the HA spike.

There is only one surface glycoprotein in influenza C, the hemagglutinin-esterase-fusion protein (HEF). Influenza C virus has, therefore, one fewer gene segments than influenza A. HEF has receptor-binding (hemaggluti-nation), fusion, and receptor-destroying activities. The receptor is sialic acid, but the activity that destroys the receptor is an esterase activity. The esterase does not remove sialic acid from proteins as does NA of influenza A. Instead it removes the 9-O-acetyl group from 9-O-acetyl-N-acetylneuraminic acid, the receptor used by influenza C, and the virus does not bind to the deacylated sialic acid.

Replication of Influenza RNA and Synthesis of mRNAs

Synthesis of influenza virus RNAs occurs in the nucleus, rather than in the cytoplasm as for most RNA viruses. This makes possible the differential splicing observed for two of the mRNAs. Following infection by the virus, the viral RNPs are transported to the nucleus and mRNA synthesis begins. During synthesis of mRNA, influenza engages in a process called "cap-snatching." Capped cellular pre-mRNAs present in the nucleus are bound by NS1, and the 5'-terminal 10-13 nucleotides, containing the 5' cap, are removed by PB2. This oligonu-cleotide is used to prime synthesis of mRNA from the influenza genome segments, as illustrated in Fig. 4.13. Once initiated, other aspects of mRNA synthesis resemble those that occur in rhabdo- and paramyxoviruses. Synthesis continues to near the end of the genome segment, where an oligo(U) stretch is encountered. Here the enzyme stutters to produce a poly(A) tail on the messenger and then releases it. In addition to its role as a primer, using a cap derived from cellular mRNA relieves the virus of the necessity of encoding enzymes required for capping and ensures that the virus mRNA has a cap suitable for the cell in which it is replicating. This mechanism also results in interference with the synthesis and transport of host mRNAs. Furthermore, because the mRNAs have a different 5' end and lack the 3' end of the antigenomic RNA, they lack promoters required for replication and packaging and are therefore dedicated mRNAs.

Each genome segment gives rise to one primary mRNA species. However, two of these can be spliced, and both the unspliced and spliced RNAs serve as messengers. Thus, two mRNAs are formed from each of two of the segments, and in total, 10 mRNAs are formed and 10 proteins are produced. The formation of the two mRNAs from segment 7 and their translation into proteins is illustrated schematically in Fig. 4.14.

When sufficient amounts of viral proteins have been synthesized and transported to the nucleus, viral RNA replication begins. Replication requires encapsidation of progeny genomic and antigenomic RNAs as described for other (-)RNA viruses, and the mechanisms that lead to a switch between synthesis of mRNAs and replication are thought to be similar to those that occur in rhabdoviruses and paramyxoviruses. During replication, the viral genome is copied into a faithful antigenomic RNA (vcRNA)

Minus-Strand RNA Viruses

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