title: Cleavage of bovine adenovirus type 3 non-structural 100K protein by protease is required for nuclear localization in infected cells but is not essential for virus replication.
abstract: The L6 region of bovine adenovirus type 3 (BAdV-3) encodes a non-structural protein named 100K. Rabbit antiserum raised against BAdV-3 100K recognized a protein of 130 kDa at 12-24 h and proteins of 130, 100, 95 and 15 kDa at 36-48 h after BAdV-3 infection. The 100K species localized to the nucleus and the cytoplasm of BAdV-3-infected cells. In contrast, 100K localized predominantly to the cytoplasm of the transfected cells. However, BAdV-3 infection of cells transfected with 100K-enhanced yellow fluorescent protein-expressing plasmid detected fluorescent protein in the nucleus of the cells, suggesting that other viral proteins may be required for the nuclear localization of 100K. Interaction of BAdV-3 100K with BAdV-3 33K protein did not alter the cytoplasmic localization of 100K. However, co-expression of BAdV-3 100K and BAdV-3 protease localized 100K to the nucleolus of the transfected cells. Subsequent analysis suggested that BAdV-3 protease cleaves 100K at two identified potential protease cleavage sites (aa 740-745 and 781-786) in transfected or BAdV-3-infected cells. The cleaved C terminus (107 aa) was localized to the nucleolus of the transfected cells. Further analysis suggested that the cleaved C terminus contains a bipartite nuclear localization signal and utilizes import receptor importin-α3 of the classical importin-α/β transport pathway for nuclear transport. Successful isolation of recombinant BAdV-3 expressing mutant 100K (substitution of alanine for glycine in the potential protease cleavage site) suggested that cytoplasmic cleavage of BAdV-3 100K by adenoviral protease is not essential for virus replication.
authors.affiliation: VIDO-InterVac, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.|Veterinary Microbiology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.
authors.forename: Nirajkumar
authors.initials: N
authors.lastname: Makadiya
authors.affiliation: VIDO-InterVac, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.|Veterinary Microbiology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.
authors.forename: Amit
authors.initials: A
authors.lastname: Gaba
authors.affiliation: Veterinary Microbiology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.|VIDO-InterVac, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.|Vaccinology & Immunotherapeutics program, School of Public Health, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E3 Canada.
authors.forename: Suresh K
authors.initials: SK
authors.lastname: Tikoo
mesh_terms: D000257:Adenoviridae Infections / Q000662:veterinary* / Q000821:virology; D025662:Adenoviruses, Porcine / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D000818:Animals; D002417:Cattle; D002418:Cattle Diseases / Q000821:virology*; D002460:Cell Line; D002466:Cell Nucleolus / Q000821:virology*; D010447:Peptide Hydrolases / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D017361:Viral Nonstructural Proteins; D010447:Peptide Hydrolases