Nuclear localization and intramolecular cleavage of N-terminally deleted NS5A protein of hepatitis C virus.
The full-size NS5A (NS5A-F) of hepatitis C virus is localized in the cytoplasm despite the presence of a functional nuclear localization signal (NLS) in its C-terminal region (amino acids (aa) 354-362). In the present study, we demonstrated that a short stretch of sequence near the N-terminus of NS5A (aa 27-38) masked the functional NLS, preventing NS5A from being transported to the nucleus. This sequence, referred to as an NLS-masking sequence, was distinct from a nuclear export signal, as it did not actively target a protein to the cytoplasm. We also found that other sequences located at either an N- (aa 1-21) or a C-terminal region (aa 353-447) were responsible for targeting NS5A to the cytoplasm. Western blot analysis of the transfected cells revealed that NS5A mutants that had been N-terminally deleted by 66 aa or more were cleaved at a certain cleavage site, generating a common fragment of ca. 40 kDa. This result implies the possible presence of a cleavage site in the NS5A sequence around aa 150, which is exposed through conformational alteration upon the N-terminal deletions.
Song
J
J
Department of Microbiology, Kobe University School of Medicine, 7-5-1 Kusunoki-cho, Chuo-ku, 650-0017, Kobe, Japan.
Nagano-Fujii
M
M
Wang
F
F
Florese
R
R
Fujita
T
T
Ishido
S
S
Hotta
H
H
D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000821:virology; D003593:Cytoplasm / Q000821:virology; D019084:Fluorescent Antibody Technique, Indirect; D016174:Hepacivirus / Q000737:chemistry / Q000235:genetics*; D051379:Mice; D017354:Point Mutation; D014170:Transformation, Genetic; D017361:Viral Nonstructural Proteins / Q000032:analysis / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase
D017361:Viral Nonstructural Proteins; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase