Accurate Measurement of the Effects of All Amino-Acid Mutations on Influenza Hemagglutinin
Abstract
:1. Introduction
2. Results
2.1. A Helper-Virus Enables Efficient Production of Mutant Virus Libraries from Plasmids
2.2. Low MOI Passage Combined with Barcoded-Subamplicon Sequencing Reveals Strong Selection against Non-Functional HA Variants
2.3. The Mutant Virus Libraries Have Reduced Bottlenecking and Yield Reproducible Measurements of Mutational Effects
2.4. The Measurements Better Reflect the Constraints on HA Evolution in Nature
2.5. A Handful of Sites Are under Very Different Selection in Our Experiments Than in Nature
2.6. Antigenic Sites in HA’s Globular Head Are Highly Tolerant of Mutations, but Stalk Epitopes Targeted by Broadly Neutralizing Antibodies Are Not
3. Discussion
4. Materials and Methods
4.1. Growth of HA-Deficient Helper Virus in HA-Expressing Cells
4.2. HA Plasmid Mutant Libraries
4.3. Generation of Mutant HA Virus Libraries from Mutant Plasmids and Helper Viruses
4.4. Barcoded Subamplicon Sequencing
4.5. Inference of Amino-Acid Preferences from Sequencing Data
4.6. Phylogenetic Modeling Using Amino-Acid Preferences
4.7. Statistical Tests
4.8. Availability of Data and Computer Code
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
HA | hemagglutinin |
MOI | multiplicity of infection |
TCID50 | 50% tissue culture infectious dose |
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Model | ΔAIC | Log Likelihood | Parameters (Optimized + Empirical): Optimized Values |
---|---|---|---|
new data + old data | 0.0 | −14933.5 | 6 (6 + 0): β = 1.82, ω = 0.51, κ = 4.95, = 0.40, = 0.18, = 0.20 |
new data | 197.6 | −15032.3 | 6 (6 + 0): β = 1.80, ω = 0.46, κ = 5.06, = 0.40, = 0.18, = 0.20 |
old data | 341.2 | −15104.1 | 6 (6 + 0): β = 1.40, ω = 0.46, κ = 4.90, = 0.39, = 0.18, = 0.20 |
Goldman–Yang M8 | 2156.8 | −16003.9 | 14 (5 + 9): = 0.01, = 1.91, = 0.02, = 0.76, κ = 4.94 |
new data + old data, averaged across sites | 2971.6 | −16419.3 | 6 (6 + 0): β = 0.50, ω = 0.20, κ = 5.38, = 0.38, = 0.18, = 0.21 |
Goldman–Yang M0 | 2980.8 | −16418.9 | 11 (2 + 9): ω = 0.19, κ = 4.88 |
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Doud, M.B.; Bloom, J.D. Accurate Measurement of the Effects of All Amino-Acid Mutations on Influenza Hemagglutinin. Viruses 2016, 8, 155. https://doi.org/10.3390/v8060155
Doud MB, Bloom JD. Accurate Measurement of the Effects of All Amino-Acid Mutations on Influenza Hemagglutinin. Viruses. 2016; 8(6):155. https://doi.org/10.3390/v8060155
Chicago/Turabian StyleDoud, Michael B., and Jesse D. Bloom. 2016. "Accurate Measurement of the Effects of All Amino-Acid Mutations on Influenza Hemagglutinin" Viruses 8, no. 6: 155. https://doi.org/10.3390/v8060155
APA StyleDoud, M. B., & Bloom, J. D. (2016). Accurate Measurement of the Effects of All Amino-Acid Mutations on Influenza Hemagglutinin. Viruses, 8(6), 155. https://doi.org/10.3390/v8060155