title: Vaccinia virus protein complex F12/E2 interacts with kinesin light chain isoform 2 to engage the kinesin-1 motor complex.
abstract: During vaccinia virus morphogenesis, intracellular mature virus (IMV) particles are wrapped by a double lipid bilayer to form triple enveloped virions called intracellular enveloped virus (IEV). IEV are then transported to the cell surface where the outer IEV membrane fuses with the cell membrane to expose a double enveloped virion outside the cell. The F12, E2 and A36 proteins are involved in transport of IEVs to the cell surface. Deletion of the F12L or E2L genes causes a severe inhibition of IEV transport and a tiny plaque size. Deletion of the A36R gene leads to a smaller reduction in plaque size and less severe inhibition of IEV egress. The A36 protein is present in the outer membrane of IEVs, and over-expressed fragments of this protein interact with kinesin light chain (KLC). However, no interaction of F12 or E2 with the kinesin complex has been reported hitherto. Here the F12/E2 complex is shown to associate with kinesin-1 through an interaction of E2 with the C-terminal tail of KLC isoform 2, which varies considerably between different KLC isoforms. siRNA-mediated knockdown of KLC isoform 1 increased IEV transport to the cell surface and virus plaque size, suggesting interaction with KLC isoform 1 is somehow inhibitory of IEV transport. In contrast, knockdown of KLC isoform 2 did not affect IEV egress or plaque formation, indicating redundancy in virion egress pathways. Lastly, the enhancement of plaque size resulting from loss of KLC isoform 1 was abrogated by removal of KLC isoforms 1 and 2 simultaneously. These observations suggest redundancy in the mechanisms used for IEV egress, with involvement of KLC isoforms 1 and 2, and provide evidence of interaction of F12/E2 complex with the kinesin-1 complex.
authors.affiliation: Department of Pathology, University of Cambridge, Cambridge, United Kingdom.
authors.forename: David C J
authors.initials: DC
authors.lastname: Carpentier
authors.affiliation: Department of Pathology, University of Cambridge, Cambridge, United Kingdom.
authors.forename: William N D
authors.initials: WN
authors.lastname: Gao
authors.affiliation: Department of Pathology, University of Cambridge, Cambridge, United Kingdom.
authors.forename: Helen
authors.initials: H
authors.lastname: Ewles
authors.affiliation: Department of Virology, Imperial College London, St. Mary's Campus, London, United Kingdom.
authors.forename: Gareth W
authors.initials: GW
authors.lastname: Morgan
authors.affiliation: Department of Pathology, University of Cambridge, Cambridge, United Kingdom.
authors.forename: Geoffrey L
authors.initials: GL
authors.lastname: Smith
mesh_terms: D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D016547:Kinesins; D018613:Microscopy, Confocal; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D014162:Transfection; D014616:Vaccinia virus / Q000378:metabolism* / Q000472:pathogenicity*; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D008869:Microtubule-Associated Proteins; D014764:Viral Proteins; D016547:Kinesins