The structurally disordered paramyxovirus nucleocapsid protein tail domain is a regulator of the mRNA transcription gradient.
The paramyxovirus RNA-dependent RNA-polymerase (RdRp) complex loads onto the nucleocapsid protein (N)-encapsidated viral N:RNA genome for RNA synthesis. Binding of the RdRp of measles virus (MeV), a paramyxovirus archetype, is mediated through interaction with a molecular recognition element (MoRE) located near the end of the carboxyl-terminal Ntail domain. The structurally disordered central Ntail section is thought to add positional flexibility to MoRE, but the functional importance of this Ntail region for RNA polymerization is unclear. To address this question, we dissected functional elements of Ntail by relocating MoRE into the RNA-encapsidating Ncore domain. Linker-scanning mutagenesis identified a microdomain in Ncore that tolerates insertions. MoRE relocated to Ncore supported efficient interaction with N, MoRE-deficient Ntails had a dominant-negative effect on bioactivity that was alleviated by insertion of MoRE into Ncore, and recombinant MeV encoding N with relocated MoRE grew efficiently and remained capable of mRNA editing. MoRE in Ncore also restored viability of a recombinant lacking the disordered central Ntail section, but this recombinant was temperature-sensitive, with reduced RdRp loading efficiency and a flattened transcription gradient. These results demonstrate that virus replication requires high-affinity RdRp binding sites in N:RNA, but productive RdRp binding is independent of positional flexibility of MoRE and cis-acting elements in Ntail. Rather, the disordered central Ntail section independent of the presence of MoRE in Ntail steepens the paramyxovirus transcription gradient by promoting RdRp loading and preventing the formation of nonproductive polycistronic viral mRNAs. Disordered Ntails may have evolved as a regulatory element to adjust paramyxovirus gene expression.
Institute for Biomedical Sciences, Georgia State University, Atlanta, GA 30303, USA.
Robert M
RM
Cox
Institute for Biomedical Sciences, Georgia State University, Atlanta, GA 30303, USA.
Stefanie A
SA
Krumm
Institute for Biomedical Sciences, Georgia State University, Atlanta, GA 30303, USA.
Vidhi D
VD
Thakkar
Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA.
Maximilian
M
Sohn
Institute for Biomedical Sciences, Georgia State University, Atlanta, GA 30303, USA.; Department of Pediatrics, Emory University School of Medicine, Atlanta, GA 30322, USA.
Richard K
RK
Plemper
D000818:Animals; D002522:Chlorocebus aethiops; D064267:Intrinsically Disordered Proteins / Q000235:genetics / Q000378:metabolism; D008459:Measles virus / Q000502:physiology*; D019590:Nucleocapsid Proteins / Q000235:genetics; D000072417:Protein Domains; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000096:biosynthesis / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000502:physiology*; D014779:Virus Replication / Q000502:physiology*
D064267:Intrinsically Disordered Proteins; D019590:Nucleocapsid Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D012324:RNA-Dependent RNA Polymerase
Nucleocapsid; Paramyxovirus; RNA-dependent RNA polymerase; Virus replication; measles virus