of RNA-Dependent RNA Polymerases
Three-Dimensional Structures of RNA-Dependent RNA Polymerases
Mechanisms of RNA Synthesis Initiation Capping Elongation Functions of Additional Polymerase Domains RNA Polymerase Oligomerization Template Specificity Unwinding the RNA Template Role of Cellular Proteins
Paradigms for Viral RNA Synthesis (+) Strand RNA Synthesis of Nested Subgenomic mRNAs (−) Strand RNA Ambisense RNA Double-Stranded RNA Unique Mechanisms of mRNA and Genome Synthesis of Hepatitis Delta Virus Do Ribosomes and RNA Polymerases Collide?
Origins of Diversity in RNA Virus Genomes Misincorporation of Nucleotides Segment Reassortment and RNA Recombination RNA Editing
LINKS FOR CHAPTER 6
Video: Interview with Dr. Karla Kirkegaard http://bit.ly/Virology_Kirkegaard
A swinging gate http://bit.ly/Virology_Twiv330
When a thing has been said and said well, have no scruple. Take it and copy it.
ANATOLE FRANCE
Introduction
The genomes of RNA viruses may be unimolecular or segmented; single stranded of (+), (−), or ambisense polarity; double stranded; or circular. These structurally diverse viral RNA genomes share a common requirement: they must be copied efficiently within the infected cell to provide both genomes for assembly into progeny virus particles and messenger RNAs (mRNAs) for the synthesis of viral proteins. The production of these RNA molecules is a unique process that has no parallel in the cell. The genomes of all RNA viruses except retroviruses and hepatitis delta virus encode an RNA-dependent RNA polymerase (RdRP) (Box 6.1) to catalyze the synthesis of new genomes and mRNAs.
Virus particles that contain (−) strand or double-stranded RNA genomes must contain the RdRP, because the incoming viral RNA can be neither translated nor copied by the cellular machinery. Consequently, the deproteinized genomes of (−) strand and double-stranded RNA viruses are not infectious. In contrast, viral particles containing a (+) strand RNA genome lack a viral polymerase; the deproteinized RNAs of these viruses are infectious because they are translated in cells to produce, among other viral proteins, the viral RNA polymerase. An exception is the retrovirus particle, which contains a (+) stranded RNA genome that is not translated but rather copied to DNA by reverse transcriptase (Chapter 10).
The mechanisms by which viral mRNA is made and the RNA genome is replicated in cells infected by RNA viruses appear even more diverse than the structure and organization of viral RNA genomes (Fig. 6.1). Nevertheless, each mechanism of viral RNA synthesis meets two essential requirements common to all infectious cycles: (i) during replication, the RNA genome must be copied from one end to the other with no loss of nucleotide sequence; and (ii) viral mRNAs that can be translated efficiently by the cellular protein synthetic machinery must be made.
In this chapter, we consider the mechanisms of viral RNA synthesis, the switch from mRNA production to genome replication, and the origins of genetic diversity. Much of our understanding of viral RNA synthesis comes from experiments with purified components. Because it is possible that events proceed differently in infected cells, the results of such in vitro studies are used to build models for the different steps in RNA synthesis, which must then be tested in vivo. While many models exist for each reaction, those presented in this chapter were selected because they are consistent with experimental results obtained in different laboratories or have been validated with simplified systems in cells in culture. The general principles of RNA synthesis deduced from such studies are illustrated with a few viruses as examples.
The Nature of the RNA Template
Secondary Structures in Viral RNA
RNA molecules are not simple linear chains but can form secondary structures that are important for RNA synthesis, translation, and assembly (Fig. 6.2). Structural features in RNA are identified by scanning the nucleotide sequence with software designed to fold the nucleic acid into energetically stable structures. Comparative sequence analysis can predict RNA secondary structures. For example, comparison of the RNA sequences of several related viruses might establish that the structure, but not the sequence, of a stem-loop is conserved. Direct evidence for specific RNA structures comes from experiments in which RNAs are treated with enzymes or chemicals that attack single- or double-stranded regions specifically. The results of such analyses can confirm that the predicted stem regions are base paired while loops are unpaired. Structures of RNA hairpins and pseudoknots have been determined by X-ray crystallography or nuclear magnetic resonance (Fig. 6.2C).
PRINCIPLES Synthesis of RNA from RNA templates
Viral RNA genomes must be copied to provide both genomes for assembly into progeny virus particles and mRNAs for the synthesis of viral proteins.
Viral RNA genomes may be naked in the virus particle [typically (+) strand
