Nuclear import of adenovirus DNA in vitro involves the nuclear protein import pathway and hsc70.
Adenovirus, a respiratory virus with a double-stranded DNA genome, replicates in the nuclei of mammalian cells. We have developed a cytosol-dependent in vitro assay utilizing adenovirus nucleocapsids to examine the requirements for adenovirus docking to the nuclear pore complex and for DNA import into the nucleus. Our assay reveals that adenovirus DNA import is blocked by a competitive excess of classical protein nuclear localization sequences and other inhibitors of nuclear protein import and indicates that this process is dependent on hsc70. Previous work revealed that the hexon (coat) protein of adenovirus is the only major protein on the surface of the adenovirus nucleocapsid that docks at the nuclear pore complex. This, together with our finding that in vitro nuclear import of hexon is inhibited by an excess of classical nuclear localization sequences, suggests a role for the hexon protein in adenovirus DNA import. However, recombinant transport factors that are sufficient for hexon import in permeabilized cells do not support DNA import, indicating that there are other as yet unidentified factors required for this process.
Saphire
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AC
Departments of Cell and Molecular Biology, Scripps Research Institute, La Jolla, California 92037, USA.
Guan
T
T
Schirmer
E C
EC
Nemerow
G R
GR
Gerace
L
L
D000256:Adenoviridae / Q000378:metabolism*; D000906:Antibodies / Q000494:pharmacology; D002213:Capsid / Q000378:metabolism; D036022:Capsid Proteins; D002467:Cell Nucleus / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D018840:HSP70 Heat-Shock Proteins / Q000276:immunology / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D017403:In Situ Hybridization; D008854:Microscopy, Electron; D009685:Nuclear Envelope / Q000378:metabolism / Q000648:ultrastructure; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000378:metabolism; D019251:Nucleocapsid / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism
D000906:Antibodies; D036022:Capsid Proteins; D004279:DNA, Viral; D018840:HSP70 Heat-Shock Proteins; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D011994:Recombinant Proteins; C018315:hexon capsid protein, Adenovirus