Characterization of a nuclear localization signal of canine parvovirus capsid proteins.
We investigated the abilities of synthetic peptides mimicking the potential nuclear localization signal of canine parvovirus (CPV) capsid proteins to translocate a carrier protein to the nucleus following microinjection into the cytoplasm of A72 cells. Possible nuclear localization sequences were chosen for synthesis from CPV capsid protein sequences (VP1, VP2) on the basis of the presence of clustered basic residues, which is a common theme in most of the previously identified targeting peptides. Nuclear targeting activity was found within the N-terminal residues 4-13 (PAKRARRGYK) of the VP1 capsid protein. While replacement of Arg10 with glycine did not affect the activity, replacement of Lys6, Arg7, or Arg9 with glycine abolished it. The targeting activity was found to residue in a cluster of basic residues, Lys5, Arg7, and Arg9. Nuclear import was saturated by excess of unlabelled peptide conjugates (showing that it was a receptor-mediated process). Transport into the nucleus was an energy-dependent and temperature-dependent process actively mediated by the nuclear pores and inhibited by wheat germ agglutinin.
Vihinen-Ranta
M
M
Department of Biological and Environmental Science, University of Jyväskylä, Finland. mvihinen@jyu.fi
Kakkola
L
L
Kalela
A
A
Vilja
P
P
Vuento
M
M
D000255:Adenosine Triphosphate / Q000502:physiology; D000818:Animals; D001692:Biological Transport; D002213:Capsid / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism*; D004285:Dogs; D019913:Nuclear Localization Signals; D017993:Parvovirus, Canine / Q000737:chemistry*; D013696:Temperature; D014407:Tumor Cells, Cultured; D014909:Wheat Germ Agglutinins / Q000494:pharmacology
D019913:Nuclear Localization Signals; D014909:Wheat Germ Agglutinins; D000255:Adenosine Triphosphate