Dataset Information

Effect of mismatch repair on the mutation rate of bacteriophage ?X174.

ABSTRACT: Viral mutation rates vary widely in nature, yet the mechanistic and evolutionary determinants of this variability remain unclear. Small DNA viruses mutate orders of magnitude faster than their hosts despite using host-encoded polymerases for replication, which suggests these viruses may avoid post-replicative repair. Supporting this, the genome of bacteriophage ?X174 is completely devoid of GATC sequence motifs, which are required for methyl-directed mismatch repair in Escherichia coli. Here, we show that restoration of the randomly expected number of GATC sites leads to an eightfold reduction in the rate of spontaneous mutation of the phage, without severely impairing its replicative capacity over the short term. However, the efficacy of mismatch repair in the presence of GATC sites is limited by inefficient methylation of the viral DNA. Therefore, both GATC avoidance and DNA under-methylation elevate the mutation rate of the phage relative to that of the host. We also found that the effects of GATC sites on the phage mutation rate vary extensively depending on their specific location within the phage genome. Finally, the mutation rate reduction afforded by GATC sites is fully reverted under stress conditions, which up-regulate repair pathways and expression of error-prone host polymerases such as heat and treatment with the base analog 5-fluorouracil, suggesting that access to repair renders the phage sensitive to stress-induced mutagenesis.

SUBMITTER: Pereira-Gomez M

PROVIDER: S-EPMC5014478 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Effect of mismatch repair on the mutation rate of bacteriophage ϕX174.

Pereira-Gómez Marianoel M Sanjuán Rafael R

Virus evolution 20150910 1

Viral mutation rates vary widely in nature, yet the mechanistic and evolutionary determinants of this variability remain unclear. Small DNA viruses mutate orders of magnitude faster than their hosts despite using host-encoded polymerases for replication, which suggests these viruses may avoid post-replicative repair. Supporting this, the genome of bacteriophage ϕX174 is completely devoid of GATC sequence motifs, which are required for methyl-directed mismatch repair in <i>Escherichia coli</i>. H ...[more]

PMID: 27774282

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Project description:Like many viruses, bacteriophage phi X174 packages its DNA genome into a procapsid that is assembled from structural intermediates and scaffolding proteins. The procapsid contains the structural proteins F, G and H, as well as the scaffolding proteins B and D. Provirions are formed by packaging of DNA together with the small internal J proteins, while losing at least some of the B scaffolding proteins. Eventually, loss of the D scaffolding proteins and the remaining B proteins leads to the formation of mature virions.phi X174 108S 'procapsids' have been purified in milligram quantities by removing 114S (mature virion) and 70S (abortive capsid) particles from crude lysates by differential precipitation with polyethylene glycol. 132S 'provirions' were purified on sucrose gradients in the presence of EDTA. Cryo-electron microscopy (cryo-EM) was used to obtain reconstructions of procapsids and provirions. Although these are very similar to each other, their structures differ greatly from that of the virion. The F and G proteins, whose atomic structures in virions were previously determined from X-ray crystallography, were fitted into the cryo-EM reconstructions. This showed that the pentamer of G proteins on each five-fold vertex changes its conformation only slightly during DNA packaging and maturation, whereas major tertiary and quaternary structural changes occur in the F protein. The procapsids and provirions were found to contain 120 copies of the D protein arranged as tetramers on the two-fold axes. DNA might enter procapsids through one of the 30 A diameter holes on the icosahedral three-fold axes.Combining cryo-EM image reconstruction and X-ray crystallography has revealed the major conformational changes that can occur in viral assembly. The function of the scaffolding proteins may be, in part, to support weak interactions between the structural proteins in the procapsids and to cover surfaces that are subsequently required for subunit-subunit interaction in the virion. The structures presented here are, therefore, analogous to chaperone proteins complexed with folding intermediates of a substrate.

Dataset Information

Effect of mismatch repair on the mutation rate of bacteriophage ?X174.

Publications

Effect of mismatch repair on the mutation rate of bacteriophage ϕX174.

Similar Datasets

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