Martins, Claudia R. F. and Johnson, Jennifer A. and Lawrence, Diane M. and Choi, Tae-Jin and Pisi, Anna Maria and Tobin, Sara L. and Lapidus, Denise and Wagner, John D. O. and Ruzin, Steven and McDonald, Kent and Jackson, Andrew O. (1998) Sonchus Yellow Net Rhabdovirus Nuclear Viroplasms Contain Polymerase-Associated Proteins. Journal of Virology, 72 (7). pp. 5669-5679. ISSN 0022-538X. http://resolver.caltech.edu/CaltechAUTHORS:MARjvir98
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We have initiated a study of the cytopathology of nucleorhabdoviruses by analyzing the subcellular localization of sonchus yellow net virus (SYNV) genomic and antigenomic RNAs and the encoded polymerase proteins. In situ hybridizations demonstrated that the minus-strand genomic RNA sequences are restricted to the nuclei of infected cells, while the complementary plus-strand antigenomic RNA sequences are present in both the nuclei and the cytoplasm. Immunofluorescence and immunogold labeling experiments also revealed that the nucleocapsid (N) protein and phosphoprotein (M2) are primarily localized to discrete regions within the nuclei and in virus particles that accumulate in perinuclear spaces. The N protein antiserum specifically labeled the nuclear viroplasms, whereas the M2 antiserum was more generally distributed throughout the nuclei. Antibody detection also indicated that the polymerase (L) protein is present in small amounts in the viroplasm. When the N and M2 proteins were expressed individually from the heterologous potato virus X (PVX) vector, both proteins preferentially accumulated in the nuclei. In addition, viroplasm-like inclusions formed in the nuclei of cells infected with the PVX vector containing the N gene. Fusions of the carboxy terminus of beta -glucuronidase to N and M2 resulted in staining of the nuclei of infected cells following expression from the PVX vector. Deletion analyses suggested that multiple regions of the N protein contain signals that are important for nuclear localization.
|Additional Information:||Copyright © 1998, American Society for Microbiology. All rights reserved. Received 16 December 1997/Accepted 23 March 1998 We thank David Baulcombe and Jim Carrington for generously providing plasmids pPC2S and pRTL2-GUS/NIaDBam, respectively, and Simon Santa Cruz for supplying antisera raised against PVX virions used in these experiments. We also thank Paula Sicurello at the Electron Microscope Facility at UC-Berkeley, Denise Schichnes at the CNR Center for Biological Imaging at UC-Berkeley, and David Jackson at the USDA Plant Gene Expression Center for assistance and technical advice about various aspects of these experiments. Michael Goodin, Ignacio Moreno, Teresa Rubio, and Angelika Fath provided suggestions and editorial comments on the manuscript. A Conselho Nacional de Desenvolvimento Cientificio e Tecnologico Fellowship was awarded to C.R.F.M., and a National Science Foundation Visiting Professorship for Women was awarded to S.L.T. This research was supported by National Science Foundation grant DMB 94-18086 awarded to A.O.J.|
|Subject Keywords:||VESICULAR STOMATITIS-VIRUS; BUSHY STUNT VIRUS; STRAND LEADER RNA; NUCLEOCAPSID PROTEIN; PLANT RHABDOVIRUS; GENE-EXPRESSION; SEQUENCE; REPLICATION; IDENTIFICATION; LOCALIZATION|
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|Deposited On:||01 Feb 2006|
|Last Modified:||26 Dec 2012 08:45|
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