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Growth of an RNA virus in single cells reveals a broad fitness distribution.

ABSTRACT: Genetic and environmental factors will influence the growth of an RNA virus, but their relative contributions are challenging to resolve because standard culture methods mask how virus particles interact with individual host cells. Here, single particles of vesicular stomatitis virus, a prototype RNA virus, were used to infect individual BHK cells. Infected cells produced 50 to 8000 progeny virus particles, but these differences were lost upon subsequent culture, suggesting the diversity of yields reflected cell-to-cell differences rather than viral genetic variation. Cells infected at different phases of their cell cycle produced from 1400 (early S) to 8700 (G(2)M) infectious virus particles, coinciding with the middle-to-upper range of the observed distribution. Fluctuations in virus and cell compositions and noisy gene expression may also contribute to the broad distribution of virus yields. These findings take a step toward quantifying how environmental variation can impact the fitness distribution of an RNA virus.

SUBMITTER: Zhu Y

PROVIDER: S-EPMC2666790 | biostudies-literature | 2009 Mar

REPOSITORIES: biostudies-literature

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Growth of an RNA virus in single cells reveals a broad fitness distribution.

Zhu Ying Y Yongky Andrew A Yin John J

Virology 20081213 1

Genetic and environmental factors will influence the growth of an RNA virus, but their relative contributions are challenging to resolve because standard culture methods mask how virus particles interact with individual host cells. Here, single particles of vesicular stomatitis virus, a prototype RNA virus, were used to infect individual BHK cells. Infected cells produced 50 to 8000 progeny virus particles, but these differences were lost upon subsequent culture, suggesting the diversity of yiel ...[more]

PMID: 19070881

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Project description:BackgroundUnderstanding the causes and consequences of phenotypic variability is a central topic of evolutionary biology. Mutations within non-coding cis-regulatory regions are thought to be of major effect since they affect the expression of downstream genes. To address the evolutionary potential of mutations affecting such regions in RNA viruses, we explored the fitness properties of mutations affecting the 5'-untranslated region (UTR) of a prototypical member of the picorna-like superfamily, Tobacco etch virus (TEV). This 5' UTR acts as an internal ribosomal entry site (IRES) and is essential for expression of all viral genes.ResultsWe determined in vitro the folding of 5' UTR using the selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) technique. Then, we created a collection of single-nucleotide substitutions on this region and evaluated the statistical properties of their fitness effects in vivo. We found that, compared to random mutations affecting coding sequences, mutations at the 5' UTR were of weaker effect. We also created double mutants by combining pairs of these single mutations and found variation in the magnitude and sign of epistatic interactions, with an enrichment of cases of positive epistasis. A correlation exists between the magnitude of fitness effects and the size of the perturbation made in the RNA folding structure, suggesting that the larger the departure from the predicted fold, the more negative impact in viral fitness.ConclusionsEvidence that mutational fitness effects on the short 5' UTR regulatory sequence of TEV are weaker than those affecting its coding sequences have been found. Epistasis among pairs of mutations on the 5' UTR ranged between the extreme cases of synthetic lethal and compensatory. A plausible hypothesis to explain all these observations is that the interaction between the 5' UTR and the host translational machinery was shaped by natural selection to be robust to mutations, thus ensuring the homeostatic expression of viral genes even at high mutation rates.

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Growth of an RNA virus in single cells reveals a broad fitness distribution.

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Growth of an RNA virus in single cells reveals a broad fitness distribution.

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