Characterization of Signal Sequences Determining the Nuclear/Nucleolar Import and Nuclear Export of the Caprine Arthritis-Encephalitis Virus Rev Protein

• Published in: Viruses 2020-08, Vol.12 (8), p.900
• Main Authors: Labrecque, Marlène, Marchand, Claude, Archambault, Denis
• Format: Article
• Language: English
• Subjects: Active Transport, Cell Nucleus; Alanine; Amino Acid Sequence; Animals; Arginine; Arthritis; Arthritis-Encephalitis Virus, Caprine - genetics; Arthritis-Encephalitis Virus, Caprine - metabolism; Binding sites; caprine arthritis-encephalitis virus; Cattle; Cell Nucleus - metabolism; Cell Nucleus - virology; Cloning; Confocal microscopy; Cytoplasm; Encephalitis; Gene Products, rev - genetics; Gene Products, rev - metabolism; Genomes; Green fluorescent protein; Green Fluorescent Proteins; HeLa Cells; Humans; Hydrophobicity; Localization; Macrophages - virology; Microscopy; nuclear export signal (NES); Nuclear Export Signals; nuclear localization signal (NLS); Nuclear Localization Signals - metabolism; Nuclear transport; nucleolar localization signal (NoLS); Nucleoli; Protein Sorting Signals; Proteins; Rev protein; small ruminant lentivirus (SRLV); Transfection
• ISSN: 1999-4915; 1999-4915
• DOI: 10.3390/v12080900

Caprine arthritis-encephalitis virus (CAEV), a lentivirus, relies on the action of the Rev protein for its replication. The CAEV Rev fulfills its function by allowing the nuclear exportation of partially spliced or unspliced viral mRNAs. In this study, we characterized the nuclear and nucleolar localization signals (NLS and NoLS, respectively) and the nuclear export signal (NES) of the CAEV Rev protein. These signals are key actors in the nucleocytoplasmic shuttling of a lentiviral Rev protein. Several deletion and alanine substitution mutants were generated from a plasmid encoding the CAEV Rev wild-type protein that was fused to the enhanced green fluorescent protein (EGFP). Following cell transfection, images were captured by confocal microscopy and the fluorescence was quantified in the different cell compartments. The results showed that the NLS region is localized between amino acids (aa) 59 to 75, has a monopartite-like structure and is exclusively composed of arginine residues. The NoLS was found to be partially associated with the NLS. Finally, the CAEV Rev protein's NES mapped between aa 89 to 101, with an aa spacing between the hydrophobic residues that was found to be unconventional as compared to that of other retroviral Rev/Rev-like proteins.