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Characterization of Filovirus Protein–Protein Interactions in Mammalian Cells Using Bimolecular Complementation
- Publication Year :
- 2011
- Publisher :
- Oxford University Press, 2011.
-
Abstract
- The virion protein 40 (VP40) and nucleoprotein (NP) of Ebola (EBOV) and Marburg viruses (MARV) play key roles during virion assembly and egress. The ability to detect interactions between VP40-VP40, VP40-NP, and NP-NP and follow these complexes as they traffic through mammalian cells would enhance our understanding of the molecular events leading to filovirus assembly and budding, and provide new insights into filovirus replication and pathogenesis. Here, we successfully employed a bimolecular complementation (BiMC) approach to visualize interactions between EBOV and MARV VP40-VP40, NP-NP, and VP40-NP proteins and localize these protein complexes in mammalian cells using confocal microscopy. We demonstrate that VP40-VP40 complexes localized predominantly at the plasma membrane, whereas VP40-NP and NP-NP complexes displayed a more dispersed pattern throughout the cytoplasm. As expected based on previous findings, efficient interactions between EBOV or MARV VP40-VP40 proteins were independent of L-domains PTAPPEY and PPPY, respectively. In contrast, the formation of EBOV or MARV VP40-VP40 complexes was dependent on the previously characterized LPLGVA and LPLGIM motifs of EBOV and MARV VP40 proteins, respectively, indicating that these motifs are important for VP40 oligomerization and subsequent budding. These results highlight the feasibility and usefulness of the BiMC approach as a strategy to further characterize both filovirus protein interactions as well as filovirus-host interactions in real time in the natural environment of the cell.
- Subjects :
- Gene Expression Regulation, Viral
Biology
medicine.disease_cause
Protein–protein interaction
Cell membrane
Viral Matrix Proteins
VP40
medicine
Immunology and Allergy
Viral Biology
Humans
Ebola virus
Genetic Complementation Test
Ebolavirus
Virology
Recombinant Proteins
Cell biology
Nucleoprotein
Complementation
Infectious Diseases
medicine.anatomical_structure
HEK293 Cells
Nucleoproteins
Marburgvirus
Cytoplasm
Virion assembly
Mutation
Subjects
Details
- Language :
- English
- Database :
- OpenAIRE
- Accession number :
- edsair.doi.dedup.....94a4172bcabc74ceadb2cb4d141e74c2