title: A functional heat shock protein 90 chaperone is essential for efficient flock house virus RNA polymerase synthesis in Drosophila cells.
abstract: The molecular chaperone heat shock protein 90 (Hsp90) is involved in multiple cellular processes including protein maturation, complex assembly and disassembly, and intracellular transport. We have recently shown that a disruption of Hsp90 activity in cultured Drosophila melanogaster cells suppresses Flock House virus (FHV) replication and the accumulation of protein A, the FHV RNA-dependent RNA polymerase. In the present study, we investigated whether the defect in FHV RNA polymerase accumulation induced by Hsp90 suppression was secondary to an effect on protein A synthesis, degradation, or intracellular membrane association. Treatment with the Hsp90-specific inhibitor geldanamycin selectively reduced FHV RNA polymerase synthesis by 80% in Drosophila S2 cells stably transfected with an inducible protein A expression plasmid. The suppressive effect of geldanamycin on protein A synthesis was not attenuated by proteasome inhibition, nor was it sensitive to changes in either the mRNA untranslated regions or protein A intracellular membrane localization. Furthermore, geldanamycin did not promote premature protein A degradation, nor did it alter the extremely rapid kinetics of protein A membrane association. These results identify a novel role for Hsp90 in facilitating viral RNA polymerase synthesis in Drosophila cells and suggest that FHV subverts normal cellular pathways to assemble functional replication complexes.
authors.affiliation: Department of Internal Medicine, Division of Infectious Diseases, 1150 W. Medical Center Drive, Ann Arbor, MI 48109-0640, USA.
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: Castorena
authors.forename: Spencer A
authors.initials: SA
authors.lastname: Weeks
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Stapleford
authors.forename: Amy M
authors.initials: AM
authors.lastname: Cadwallader
authors.forename: David J
authors.initials: DJ
authors.lastname: Miller
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016227:Benzoquinones / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004331:Drosophila melanogaster / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D047029:Lactams, Macrocyclic / Q000494:pharmacology; D008969:Molecular Sequence Data; D011132:Polyribosomes / Q000378:metabolism; D012328:RNA Viruses / Q000201:enzymology; D012333:RNA, Messenger / Q000378:metabolism; D012367:RNA, Viral / Q000378:metabolism; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase; C001277:geldanamycin