title: Nuclear import of the respiratory syncytial virus matrix protein is mediated by importin beta1 independent of importin alpha.
abstract: The matrix (M) protein of respiratory syncytial virus (RSV) plays an important role in virus assembly through specific interactions with RSV nucleocapsids and envelope glycoproteins in the cytoplasm as well as with the host cell membrane. We have previously shown that M localizes to the nucleus of infected cells at an early stage in the RSV infection cycle, where it may be instrumental in inhibiting host cell processes. The present study uses transient expression of M as well as a truncated green fluorescent protein (GFP) fusion derivative to show for the first time that M is able to localize in the nucleus in the absence of other RSV gene products, through the action of amino acids 110-183, encompassing the nucleic acid binding regions of the protein, that are sufficient to target GFP to the nucleus. Using native PAGE, ELISA-based binding assays, a novel Alphascreen assay, and an in vitro nuclear transport assay, we show that M is recognized directly by the importin beta1 nuclear import receptor, which mediates its nuclear import in concert with the guanine nucleotide-binding protein Ran. Retention of M in the nucleus through binding to nuclear components, probably mediated by the putative zinc finger domain of M, also contributes to M nuclear accumulation. This is the first report of the importin binding and nuclear import properties of a gene product from a negative sense RNA virus, with implications for the function of RSV M and possibly other viral M proteins in the nucleus of infected cells.
authors.affiliation: Department of Microbiology, Monash University, Clayton, Australia.
authors.forename: Reena
authors.initials: R
authors.lastname: Ghildyal
authors.forename: Adeline
authors.initials: A
authors.lastname: Ho
authors.forename: Kylie M
authors.initials: KM
authors.lastname: Wagstaff
authors.forename: Manisha M
authors.initials: MM
authors.lastname: Dias
authors.forename: Chenoa L
authors.initials: CL
authors.lastname: Barton
authors.forename: Patricia
authors.initials: P
authors.lastname: Jans
authors.forename: Phillip
authors.initials: P
authors.lastname: Bardin
authors.forename: David A
authors.initials: DA
authors.lastname: Jans
mesh_terms: D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism / Q000821:virology*; D002614:Chelating Agents / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D051381:Rats; D012136:Respiratory Syncytial Viruses / Q000378:metabolism*; D014763:Viral Matrix Proteins / Q000737:chemistry / Q000378:metabolism*; D028901:alpha Karyopherins / Q000378:metabolism*; D028961:beta Karyopherins / Q000378:metabolism*; D020931:ran GTP-Binding Protein / Q000378:metabolism
chemical_list: D002614:Chelating Agents; C424166:KPNB1 protein, human; C094999:Kpnb1 protein, mouse; D009687:Nuclear Proteins; D014763:Viral Matrix Proteins; D028901:alpha Karyopherins; D028961:beta Karyopherins; D004492:Edetic Acid; D020931:ran GTP-Binding Protein