Functional Dissection of the Dominant Role of CD55 in Protecting Vesicular Stomatitis Virus against Complement-Mediated Neutralization
Abstract
1. Introduction
2. Materials and Methods
2.1. Statement on Biosafety Clearance
2.2. Cell Lines and Virus
2.3. Antibodies
2.4. Western Blotting
2.5. Taqman Real-Time PCR
2.6. Cycloheximide (CHX) Chase Assay
2.7. Flow Cytometry
2.8. ELISA
2.9. Purification of VSV and Western Blotting
2.10. Virus Neutralization Assay
3. Results
3.1. Differential Expression of Complement Regulatory Proteins CD55 and CD46 in VSV-Infected Cells
3.2. The Cell Surface Expression of CD55 Is Greater Than That of CD46 on VSV-Infected Cells
3.3. VSV Down-Regulates the Expression of CD46 and CD55 mRNAs
3.4. CD55 Is More Stable Than CD46 in HeLa Cells
3.5. CD46 and CD55 Incorporation into the VSV Envelope Is Independent of the Time Period of Virus Egress
3.6. Cell Type Differences in the Protection of VSV against Complement
4. Discussion
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Gene | Assay ID | Accession No. | Amplicon Size (bp) |
---|---|---|---|
CD55 | Hs00892618_m1 | 1604 | 98 |
CD46 | Hs00611257_m1 | 4179 | 148 |
18s rRNA | Hs99999901_s1 | HSRRN18S | 187 |
Time Range of Harvest (h) | CD55 (ng/5 μg Virus) a | CD46 (ng/5 μg Virus) b | Normalized VSV-G Levels against the 6–12 h Levels c | Ratio of CD55/G d | Ratio of CD46/G e | Normalized VSV-M Levels against the 6–12 h Levels f | Ratio of CD55/M g | Ratio of CD46/M h |
---|---|---|---|---|---|---|---|---|
6-12 | 50 | 2.14 | 1 | 50 | 2.14 | 1 | 50 | 2.14 |
12-18 | 37.45 | 1.74 | 0.76 | 49.3 | 2.29 | 0.78 | 48.0 | 2.23 |
18-24 | 17.4 | 1.29 | 0.43 | 40.5 | 3.0 | 0.37 | 47.0 | 3.49 |
Serial Number | Time Range of Harvest (h) | VSV-HeLa Fold Reduction (VSV+PBS/VSV+NHS) | VSV-A549 Fold Reduction (VSV+PBS/VSV+NHS) |
---|---|---|---|
1. | 0–6 | 13.75 | 56.8 |
2. | 6–12 | 10 | 236.36 |
3. | 12–18 | 80 | 2823.52 |
4. | 18–24 | 76.19 | 450 |
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Asok Kumar, N.; Muraleedharan Suma, S.; Kunnakkadan, U.; Nag, J.; Koolaparambil Mukesh, R.; Lyles, D.S.; Johnson, J.B. Functional Dissection of the Dominant Role of CD55 in Protecting Vesicular Stomatitis Virus against Complement-Mediated Neutralization. Viruses 2021, 13, 373. https://doi.org/10.3390/v13030373
Asok Kumar N, Muraleedharan Suma S, Kunnakkadan U, Nag J, Koolaparambil Mukesh R, Lyles DS, Johnson JB. Functional Dissection of the Dominant Role of CD55 in Protecting Vesicular Stomatitis Virus against Complement-Mediated Neutralization. Viruses. 2021; 13(3):373. https://doi.org/10.3390/v13030373
Chicago/Turabian StyleAsok Kumar, Nisha, Sreenath Muraleedharan Suma, Umerali Kunnakkadan, Joydeep Nag, Reshma Koolaparambil Mukesh, Douglas S. Lyles, and John Bernet Johnson. 2021. "Functional Dissection of the Dominant Role of CD55 in Protecting Vesicular Stomatitis Virus against Complement-Mediated Neutralization" Viruses 13, no. 3: 373. https://doi.org/10.3390/v13030373
APA StyleAsok Kumar, N., Muraleedharan Suma, S., Kunnakkadan, U., Nag, J., Koolaparambil Mukesh, R., Lyles, D. S., & Johnson, J. B. (2021). Functional Dissection of the Dominant Role of CD55 in Protecting Vesicular Stomatitis Virus against Complement-Mediated Neutralization. Viruses, 13(3), 373. https://doi.org/10.3390/v13030373