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Measles virus nucleocapsid protein can function in Sendai virus defective interfering particle genome synthesis in
R Chandrika1, T Myers, S A Moyer
1Department of Molecular Genetics and Microbiology, University of Florida College of Medicine, Gainesville 32610.
Abstract:
The Sendai virus P and L proteins, the viral RNA polymerase, and the nucleocapsid protein, NP, synthesized in a transient mammalian expression system support the replication of Sendai virus defective interfering particle (DI) genome RNA in vitro. We have shown that the measles virus nucleocapsid protein, N, can substitute for the Sendai NP protein in genome synthesis. The chimeric product nucleocapsids, which contained Sendai RNA encapsidated with measles N protein, were atypical since they were sensitive to micrococcal nuclease digestion, unlike wild-type Sendai or measles nucleocapsids. The utilization of measles N protein required the endogenous Sendai virus RNA polymerase, since DI nucleocapsids free of polymerase were not replicated. Although both Sendai virus NP and P proteins and measles N and P proteins formed complexes when they were coexpressed, sedimentation analysis showed that measles N protein self-assembled and did not form a complex when expressed with the Sendai P protein. Furthermore, when the Sendai P-L polymerase complex was provided separately, measles N protein alone synthesized DI genome RNA in the absence of Sendai P protein. These data suggest that the self-assembled form of measles N protein functions in Sendai DI genome synthesis.
Insights
Measles virus N protein can substitute for Sendai virus NP protein in RNA replication. Self-assembled measles N protein functions in Sendai DI genome synthesis, even without Sendai P protein.
Area of Science:
- Virology
- Molecular Biology
- Protein Interactions
Background:
- Sendai virus RNA replication requires viral P and L proteins (RNA polymerase) and nucleocapsid protein (NP).
- Defective interfering (DI) particles are viral genomes that require helper virus proteins for replication.
- Understanding protein substitution and function is crucial for viral replication mechanisms.
Purpose of the Study:
- To investigate if measles virus nucleocapsid protein (N) can substitute for Sendai virus NP in DI genome RNA replication.
- To determine the role of Sendai virus RNA polymerase and protein interactions in this process.
- To elucidate the functional form of measles N protein in viral RNA synthesis.
Main Methods:
- Transient mammalian expression system to synthesize viral proteins.
- In vitro replication assays using Sendai virus DI genome RNA.
- Micrococcal nuclease digestion to assess nucleocapsid structure.
- Co-expression and sedimentation analysis to study protein complex formation.
Main Results:
- Measles virus N protein successfully substituted for Sendai NP protein in DI genome RNA replication.
- Chimeric nucleocapsids (Sendai RNA with measles N) were sensitive to nuclease digestion.
- Measles N protein-mediated replication required Sendai virus RNA polymerase.
- Measles N protein self-assembled and could synthesize RNA independently of Sendai P protein.
Conclusions:
- Measles virus N protein can functionally substitute for Sendai virus NP in DI genome RNA synthesis.
- The self-assembled form of measles N protein is sufficient for Sendai DI genome replication when the viral RNA polymerase is present.
- This suggests distinct functional roles for NP/N and P/L proteins in paramyxovirus replication.