The Measles Virus Nucleocapsid Protein Tail Domain Is Dispensable for Viral Polymerase Recruitment and Activity

• Published in: The Journal of biological chemistry Vol. 288; no. 41; pp. 29943 - 29953
• Main Authors: Krumm, Stefanie A., Takeda, Makoto, Plemper, Richard K.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 11.10.2013; American Society for Biochemistry and Molecular Biology
• Subjects: Animal Viruses; Animals; Binding Sites - genetics; Cell Line; Chlorocebus aethiops; Immunoblotting; Measles virus - genetics; Measles virus - metabolism; Microbiology; Models, Molecular; Mutation; Nucleocapsid Proteins; Nucleoproteins - chemistry; Nucleoproteins - genetics; Nucleoproteins - metabolism; Protein Binding; Protein Structure, Tertiary; Protein Subunits - chemistry; Protein Subunits - genetics; Protein Subunits - metabolism; Ribonucleoproteins - chemistry; Ribonucleoproteins - genetics; Ribonucleoproteins - metabolism; RNA-Dependent RNA Polymerase - chemistry; RNA-Dependent RNA Polymerase - genetics; RNA-Dependent RNA Polymerase - metabolism; Vero Cells; Viral Polymerase; Viral Proteins - chemistry; Viral Proteins - genetics; Viral Proteins - metabolism; Viral Replication; Virus; Virus Replication; Virus; Viral Replication; Animal Viruses; Microbiology; Viral Polymerase
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M113.503862

Paramyxovirus genomes are ribonucleoprotein (RNP) complexes consisting of nucleoprotein (N)-encapsidated viral RNA. Measles virus (MeV) N features an amino-terminal RNA-binding core and a 125-residue tail domain, of which only the last 75 residues are considered fully mobile on the nucleocapsid surface. A molecular recognition element (MoRE) domain mediates binding of the viral phosphoprotein (P). This P N-tail interaction is considered instrumental for recruiting the polymerase complex to the template. We have engineered MeV N variants with tail truncations progressively eliminating the MoRE domain and upstream tail sections. Confirming previous reports, RNPs with N truncations lacking the carboxyl-terminal 43-residues harboring the MoRE domain cannot serve as polymerase template. Remarkably, further removal of all tail residues predicted to be surface-exposed significantly restores RNP bioactivity. Insertion of structurally dominant tags into the central N-tail section reduces bioactivity, but the negative regulatory effect of exposed N-tail stems is sequence-independent. Bioactive nucleocapsids lacking exposed N-tail sections are unable to sustain virus replication, because of weakened interaction of the advancing polymerase complex with the template. Deletion of the N-MoRE-binding domain in P abrogates polymerase recruitment to standard nucleocapsids, but polymerase activity is partially restored when N-tail truncated RNPs serve as template. Revising central elements of the current replication model, these data reveal that MeV polymerase is capable of productively docking directly to the nucleocapsid core. Dispensable for polymerase recruitment, N-MoRE binding to P-tail stabilizes the advancing polymerase-RNP complex and may rearrange unstructured central tail sections to facilitate polymerase access to the template. Background: The carboxyl-terminal tail domain of the paramyxovirus nucleoprotein is considered instrumental for polymerase recruitment to the template. Results: Truncated nucleoproteins reveal that the tail domain is dispensable for viral polymerase loading and activity. Conclusion: The viral polymerase complex is capable of productively docking directly to the nucleocapsid core. Significance: This finding alters the current paradigm of paramyxovirus polymerase recruitment.