Arch Virol (2004) 149: 1783–1799
Characterization and nuclear localization
of the ﬁber protein encoded by the late
region 7 of bovine adenovirus type 3
Q. Wu, Y. Chen, V. Kulshreshtha, and S. K. Tikoo
Vaccine and Infectious Disease Organization,
University of Saskatchewan, Saskatoon, Canada
Received November 26, 2003; accepted February 10, 2004
Published online April 15, 2004
Summary. To identify the protein encoded by the L7 region of bovine adenovirus-
3 (BAdV-3), speciﬁc antisera were raised by immunizing rabbits with bacterial
fusion proteins encoding the N-terminus or C-terminus of the BAdV-3 ﬁber
protein. Immunoprecipitation and Western blot analysis conﬁrmed that the ﬁber is
expressed as a 102 kDa glycoprotein, which is localized to the nucleus of infected
cells. To identify the nuclear localization signals (NLS), BAdV-3 ﬁber deletion mu-
tants and GFP/β-galactosidase fusion proteins were expressed in transfected cells,
and subcellular localization was visualized by immunoﬂuorescence microscopy.
Analysis of deletion mutants localized the NLS to the N-terminal 41 amino acids.
Analysis of the N-terminal 41 amino acids identiﬁed a cluster of basic residues
between amino acid 14 and 20. Substitution of the basic residues (
with acidic residues (
) resulted in the accumulation of ﬁber in the
were not sufﬁcient for
efﬁcient transport of a cytoplasmic protein GFP/β-galactosidase to the nucleus.
The recombinant BAdV-3 expressing mutant ﬁber containing
was unable to replicate efﬁciently in Madin-Darby bovine kidney
cells, suggesting that the NLS of ﬁber carries out important in vivo functions.
The adenovirus (AdV) ﬁber is a structural protein, which forms antenna-like
projections extending from the vertices of icosahedral virus capsid . All known
mammalian AdVs have one ﬁber per vertex, noncovalently attached to a penton
base protein . However, avian AdVs have two ﬁbers per vertex . The AdV
Published as VIDO journal series no. 359.