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Chapter 17 : Coding Biases and Viral Fitness
Category: Viruses and Viral Pathogenesis
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Codon usage bias has also been described for DNA and RNA viruses. Among the latter, poliovirus (PV) has selected through a codon bias similar to that of its human host species (“optimized”), while the codon bias of hepatitis A virus (HAV) is very different from that of its host (“deoptimized”). Picornavirus internal ribosome entry site (IRES) types have probably evolved by gradual addition of domains and elements that improved their function in ribosome recruitment or otherwise conferred regulation to the process of viral protein synthesis in a specific cell environment. The highly inefficient IRES combined with the lack of a mechanism to induce cellular shutoff leads in HAV to an unfair competition for the cellular translational machinery. An intriguing connection exists between codon bias, codon pair bias, and dinucleotide bias in mammalian genomes. Viral genomes, especially of RNA viruses and retroviruses, are short enough to make them amenable to whole-genome synthesis with currently available technology. Such freedom of design can provide tremendous power to reengineer DNA- and RNA-coding sequences at will to study the impact on viral fitness of large-scale changes in codon bias, codon pair bias, dinucleotide biases, GC content, RNA secondary structures, and other sequence signatures, with the aim to develop a new platform for vaccine design and genetic engineering. The codon usage selected through evolution by PV, HAV and its contribution to their in vivo fitness are still not completely elucidated, but it is certainly remarkable how they follow clearly different strategies.
Fitness loss of HAV and fitness gain of PV as a response to ActD-induced cellular shutoff. Viruses produced in the presence of 0.05 and 0.2 μg of ActD/ml in comparison with virus produced in the absence of the drug are depicted. The mean titer of 11 virus passages in the absence of the drug was given an arbitrary value of 100. The mean titer of 11 passages of the viruses propagated in the presence of ActD is expressed as the percentage of production in the absence of the drug.
Fitness loss followed by fitness recovery during the processes of HAV adaptation to different ActD concentrations. Infectious HAV titer production per cell is shown during the first 20 adaptation passages from 0.00 to 0.05, from 0.05 to 0.0, from 0.05 to 0.2, and from 0.2 to 0.05 μg of ActD/ml. TCID50, 50% tissue culture infective dose.
Growth competition experiments showing the strong virus adaptation to the host microenvironment induced by ActD (abbreviated as AMD in the figure). (Top) Populations adapted to growth in the absence and in the presence of 0.05 μg of ActD/ml were mixed at a 1:1 ratio and grown in the presence of 0.05 μg of ActD/ml. (Bottom) Populations adapted to growth with 0.05 and 0.2 μg of ActD/ml were mixed at a 1:1 ratio and grown in the presence of 0.2 μg of ActD/ml.
A genomic analysis of populations adapted to replication in the presence of ActD shows its association with a change in codon usage in the structural protein-coding region but not in the polymerase region. The relative proportion (percentage) of the newly generated codons detected during the process of HAV adaptation to ActD is shown. Codons are sorted as being similarly frequent (black), less frequent (light gray), or more frequent (dark gray) than the original ones with respect to cell host codon usage. (A) Codon usage variation in the capsid region in the absence of ActD. (B) Codon usage variation in the capsid region in the presence of 0.05 μg of ActD/ml from passages 4 to 85 and in the presence of 0.2 μg of ActD/ml from passages 20 (20′) to 38 (38′). (C) Codon usage variation in the polymerase region in the absence of ActD. (D) Codon usage variation in the polymerase region in the presence of 0.05 μg of ActD from passages 4 to 85 and in the presence of 0.2 μg of ActD/ml from passages 20 (20′) to 38 (38′).
Capsid folding is independent of Hsp90 in HAV and dependent on Hsp90 in PV. Relative infectious rates for HAV and PV per cell in the presence of increasing concentrations of geldanamycin, an Hsp90 inhibitor ( 25 ), are shown. Viral production at each geldanamycin concentration is expressed as a percentage of viral production in the absence of the drug. IC50, 50% inhibitory concentration.
(A) Calculated CPB scores for all 14,795 annotated human genes. Each dot represents the calculated CPB score of a gene plotted against its amino acid length. Underrepresented codon pairs yield negative scores. Various PV constructs are represented by symbols: PV(M)-wt, wt PV (CPB = –0.02). Lower CPB scores result in lower translation and higher attenuation. (B) Structures of the various chimeric, partly synthetic PV constructs and their viabilities in cultured cells. Nucleotide positions in the viral genome are shown. (C) One-step growth curve with respect to PFU. A multiplicity of infection of 2 was used to infect a monolayer of HeLa R19 cells. Symbols: open squares, PV(M)-wt; solid circles, PV-Max; open diamonds, PV-Min755-1513; asterisks, PV-Min1513-2470; solid diamonds, PV-MinXY; open triangles, PV-MinZ. (D) Same experiment as in panel C, but with results graphed with respect to the number of viral particles instead of PFU. (E) Plaque phenotypes of viruses after 72 h of incubation and staining with crystal violet (plate diameter, 35 mm). (Modified from Coleman et al., 2008 [ 18 ].)
In PV, translational selection, i.e., the adaptation of codon usage to cellular codon usage and thus to the tRNA pools for a quantitatively highly efficient and accurate rate of translation, is the main driving evolutionary force of codon usage. The avoidance of certain codon pair combinations (codon pair bias) that slow down the translational rate is an additional regulation step for highly efficient translation. In contrast, in HAV, fine-tuning translation kinetics selection, i.e., the right combination of common and rare codons allowing a regulated ribosome traffic rate to ensure proper protein folding, is the main driving evolutionary force of its codon usage. Ribosome stallings at rare codon positions allow a sequential folding. When such a combination is lost due to a change in the tRNA pools (i.e., artificial shutoff induced by ActD [abbreviated as AMD in the figure]), capsid folding is altered and a readaptation of codon usage takes place to regain the kinetics of translation and proper protein folding.
Dinucleotide statistics in poliovirus, hepatitis A virus, and various synthetic poliovirus capsid coding sequences a