Kinetic insights into the mechanism of influenza A virus membrane fusion
Abstract/Contents
- Abstract
- Host lipid composition influences many stages of the influenza A virus (IAV) entry process, including: initial binding of IAV to sialylated glycans, fusion between the viral envelope and the host membrane, and the formation of a fusion pore through which the viral genome is transferred into a target cell. In particular, target membrane cholesterol has been shown to preferentially associate with virus receptors and alter physical properties of the membrane like fluidity and curvature. These properties affect both IAV binding and fusion, which makes it difficult to isolate the role of cholesterol in IAV fusion from receptor binding effects. Here, I develop a new fusion assay that uses synthetic DNA-lipid conjugates as surrogate viral receptors to tether virions to target vesicles. To avoid the possibly perturbative effect of adding self-quenched concentrations of dye-labeled lipids or dye to the viral membrane and viral contents, I tether virions to lipid- or content-labeled target vesicles, and use fluorescence microscopy to detect individual, pH-triggered IAV membrane fusion events. Through this approach, I find that cholesterol in the target membrane enhances the efficiency of IAV lipid and content mixing, while the rate of both processes is independent of cholesterol composition. I also find that the single virus kinetics of IAV lipid mixing to target membranes with different cholesterol compositions is independent of receptor binding, suggesting that cholesterol-mediated spatial clustering of viral receptors within the target membrane does not significantly affect IAV hemifusion. Finally, I show that cholesterol decreases the fraction of content mixing events that result in content loss, which provides direct experimental evidence to support the hypothesis that the negative spontaneous curvature of cholesterol stabilizes pore formation in IAV entry and limits leakage following pore formation. This approach is a promising strategy for studying the single-virus lipid and content mixing kinetics of other enveloped viruses.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2021; ©2021 |
| Publication date | 2021; 2021 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Liu, Katherine |
|---|---|
| Degree supervisor | Boxer, Steven G. (Steven George), 1947- |
| Thesis advisor | Boxer, Steven G. (Steven George), 1947- |
| Thesis advisor | Cegelski, Lynette |
| Thesis advisor | Cui, Bianxiao |
| Degree committee member | Cegelski, Lynette |
| Degree committee member | Cui, Bianxiao |
| Associated with | Stanford University, Department of Chemistry |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Katherine Naomi Liu. |
|---|---|
| Note | Submitted to the Department of Chemistry. |
| Thesis | Thesis Ph.D. Stanford University 2021. |
| Location | https://purl.stanford.edu/zr161gj3729 |
Access conditions
- Copyright
- © 2021 by Katherine Liu
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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