Thesis ; Online: Elucidation of Orthopoxvirus Adaptation Mechanisms by Proteomics and Genomics
2018
Abstract: Adaptive changes of viruses enable the virus-host coevolution and infection of new host species. While viruses with an RNA genome adapt primarily by genomic variabilities, adaptation mechanisms of viruses with a DNA genome have remained largely elusive. ... ...
| Abstract | Adaptive changes of viruses enable the virus-host coevolution and infection of new host species. While viruses with an RNA genome adapt primarily by genomic variabilities, adaptation mechanisms of viruses with a DNA genome have remained largely elusive. Since genomes of DNA viruses are more stable compared to genomes of RNA viruses, it is assumed that mechanisms other than genomic changes underlie the adaptation of DNA viruses. However, these mechanisms have hardly been investigated so far. Since isolated virus particles are capable of crossing the species barrier, it is assumed that adaptive changes can be identified in the virus particles themselves. Therefore, this study aimed to elucidate adaptation mechanisms of DNA virus particles by proteomics and genomics technologies. While next-generation sequencing technologies are frequently used to analyze genomic changes of viruses, adaptive changes of virus particles by mass spectrometry-based proteomics have not been analyzed yet. In the present study, cowpox virus (CPXV) was used as a model virus and cell culture as a model system to study adaptive changes of DNA virus particles. CPXV is a member of the genus Orthopoxvirus (OPV) and is able infect a remarkably broad range of host species, e.g. rats, cats, elephants and humans. Increasing numbers of CPXV infections in Europe underline the need for a comprehensive understanding of OPV adaptation mechanisms. CPXV particles were isolated from a rat, which is a natural host of these viruses, and serially passaged five times in a rat and a human cell line. During passaging, an increase in viral fitness was observed exclusively in human cells, suggesting an adaptation of virus particles. Strikingly, proteome analysis revealed that the composition of virus particles changed in a cell line-specific manner, while the viral genome remained overall stable during passaging in both cell lines. Because several ubiquitination sites in virus proteins were identified, the role of ubiquitin for CPXV infection was analyzed. It was shown by the first global ubiquitination site analysis of virus particles that ubiquitin is a major conserved CPXV modification. Additionally, the dependence of CPXV replication on this protein modification was verified, making ubiquitination changes an attractive hypothesis of OPV adaptation. Furthermore, it was shown that CPXV particles incorporate intact transcripts, which presumably enable the rapid expression of viral immunomodulatory proteins upon infection. Summarized, the results of the present study lead to new findings about OPV adaptation mechanisms in vitro. These mechanisms may also apply to in vivo adaptation of DNA viruses and may enable, for example, a crossing of the species barrier. The methods established in this study enable the further characterization of OPV adaptation and, moreover, can be applied to elucidate adaptation mechanisms of viruses belonging to other families. |
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| Keywords | Biomedical engineering|Bioinformatics |
| Subject code | 570 |
| Language | ENG |
| Publishing date | 2018-01-01 00:00:01.0 |
| Publisher | Technische Universitaet Berlin (Germany) |
| Publishing country | us |
| Document type | Thesis ; Online |
| Database | BASE - Bielefeld Academic Search Engine (life sciences selection) |
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