Molecular biology of the human coronaviruses, MERS-CoV and SARS-CoV
University of Bristol, 2023
Online
Hochschulschrift
Zugriff:
MERS-CoV and SARS-CoV-2 cause respiratory illnesses that may lead to severe respiratory diseases in humans. To help answer the question of why some coronaviruses are lethal to humans but not to their animal hosts, I used high-throughput quantitative proteomics to analyze the expression of proteins involved in the innate immune system response in bat, camel, and human cells. Bioinformatic analysis showed a difference in the expression levels of proteins involved in the innate immune response between the three species, especially the pro-inflammatory cytokines associated with viral diseases' severity and mortality. Due to the limited information regarding the role of MERS-CoV accessory proteins ORF4A and ORF4B in virus replication, our lab investigated MERS-CoV ORF4A and MERS-CoV-ORF4B host protein-protein interactions using high-throughput quantitative proteomics in HEK293 cells. Based on the generated data, I selected six proteins (NDH II, NF45, HNRNPAB, HNRNPC, KPNA4, and HNRNPA2B1) to validate their interactions with the MERS-CoV ORF4A or MERS-CoV ORF4B in human cells. The interactions of MERS-CoV ORF4A & ORF4B proteins with bat and camel equivalent proteins were also examined to investigate whether they interact with these viral proteins with higher or lower affinity than their human counterparts or whether these interactions are species-specific. This analysis could help understand the potential roles of MERS-CoV ORF4A & ORFB proteins in virus replication and why MERS-CoV infection is lethal for humans but not for its animal hosts. Open reading frame 10 (ORF10) is one of the unique features of the SARS-CoV-2 genome, and its potential function remains controversial. One aim of this work is to shed light on the possible role of ORF10 in SARS-CoV-2 infection. In this study, I used immunoprecipitation of FLAG-tagged ORF10 proteins to identify its cellular-interacting partners. In addition, reverse genetics techniques were used to rescue the SARS-CoV-2 virus with either the ORF10 protein knocked out or replaced with fluorescent markers. The cellular proteome of SARS-CoV-2 ORF10 mutant infected cells was investigated using high-throughput proteomics analysis to uncover more information on how ORF10 protein may be modulating the host's innate immune proteins or other aspects. Finally, the replicative fitness of the SARS-CoV-2 ORF10 mutant was characterized in different cell lines.
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Molecular biology of the human coronaviruses, MERS-CoV and SARS-CoV
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Autor/in / Beteiligte Person: | Almuqrin, Abdulaziz M. ; Davidson, Andrew ; Matthews, David |
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Veröffentlichung: | University of Bristol, 2023 |
Medientyp: | Hochschulschrift |
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