title: Role of G protein and protein kinase signalling in influenza virus budding in MDCK cells.
abstract: Recently, we have shown that influenza virus budding in MDCK cells is regulated by metabolic inhibitors of ATP and ATP analogues (Hui & Nayak, Virology 290, 329-341, 2001 ). In this report, we demonstrate that G protein signalling stimulators such as sodium fluoride, aluminium fluoride, compound 48/80 and mastoparan stimulated the budding and release of influenza virus. In contrast, G protein signalling blockers such as suramin and NF023 inhibited virus budding. Furthermore, in filter-grown lysophosphatidylcholine-permeabilized virus-infected MDCK cells, membrane-impermeable GTP analogues, such as guanosine 5'-O-(3-thiotriphosphate) or 5'-guanylylimidodiphosphate caused an increase in virus budding, which could be competitively inhibited by adding an excess of GTP. These results suggest that the G protein is involved in the regulation of influenza virus budding. We also determined the role of different protein kinases in influenza virus budding. We observed that specific inhibitors or activators of protein kinase A (H-89 and 8-bromoadenosine 3',5'-cyclic monophosphate) or of protein kinase C (bisindolylmaleimide I and Ro-32-0432) or of phosphatidylinositol 3-kinase (LY294002 and wortmannin) did not affect influenza virus budding. However, the casein kinase 2 (CK2) inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole decreased virus budding. We further observed an increase in the CK2 activity during the replication cycle of influenza virus, although Western blot analysis did not reveal any increase in the amount of CK2 protein in virus-infected cells. Also, in digitonin-permeabilized MDCK cells, the introduction of CK2 substrate peptides caused a down-regulation of virus budding. These results suggest that CK2 activity also regulates influenza virus budding.
authors.affiliation: Department of Microbiology, Immunology and Molecular Genetics, Jonsson Comprehensive Cancer Center (JCCC), UCLA School of Medicine, Los Angeles, CA 90095-1747, USA1.
authors.forename: Eric Ka-Wai
authors.initials: EK
authors.lastname: Hui
authors.affiliation: Department of Microbiology, Immunology and Molecular Genetics, Jonsson Comprehensive Cancer Center (JCCC), UCLA School of Medicine, Los Angeles, CA 90095-1747, USA1.
authors.forename: Debi P
authors.initials: DP
authors.lastname: Nayak
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000494:pharmacology; D047390:Casein Kinase II; D002460:Cell Line; D004285:Dogs; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D015967:Gene Expression Regulation, Viral; D006801:Humans; D009980:Influenza A virus / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins; D047908:Intracellular Signaling Peptides and Proteins; D010455:Peptides; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D015398:Signal Transduction; D012969:Sodium Fluoride / Q000494:pharmacology; D013498:Suramin / Q000031:analogs & derivatives* / Q000494:pharmacology; D014862:Wasp Venoms / Q000494:pharmacology; D003189:p-Methoxy-N-methylphenethylamine / Q000494:pharmacology
chemical_list: D002352:Carrier Proteins; D036341:Intercellular Signaling Peptides and Proteins; D047908:Intracellular Signaling Peptides and Proteins; C105374:NF023; D010455:Peptides; D047428:Protein Kinase Inhibitors; D014862:Wasp Venoms; C015482:protein kinase modulator; D003189:p-Methoxy-N-methylphenethylamine; D013498:Suramin; C037500:mastoparan; D012969:Sodium Fluoride; D011494:Protein Kinases; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases; D019204:GTP-Binding Proteins