title: Genome-wide RNA interference in Drosophila cells identifies G protein-coupled receptor kinase 2 as a conserved regulator of NF-kappaB signaling.
abstract: Because NF-kappaB signaling pathways are highly conserved in evolution, the fruit fly Drosophila melanogaster provides a good model to study these cascades. We carried out an RNA interference (RNAi)-based genome-wide in vitro reporter assay screen in Drosophila for components of NF-kappaB pathways. We analyzed 16,025 dsRNA-treatments and identified 10 novel NF-kappaB regulators. Of these, nine dsRNA-treatments affect primarily the Toll pathway. G protein-coupled receptor kinase (Gprk)2, CG15737/Toll pathway activation mediating protein, and u-shaped were required for normal Drosomycin response in vivo. Interaction studies revealed that Gprk2 interacts with the Drosophila IkappaB homolog Cactus, but is not required in Cactus degradation, indicating a novel mechanism for NF-kappaB regulation. Morpholino silencing of the zebrafish ortholog of Gprk2 in fish embryos caused impaired cytokine expression after Escherichia coli infection, indicating a conserved role in NF-kappaB signaling. Moreover, small interfering RNA silencing of the human ortholog GRK5 in HeLa cells impaired NF-kappaB reporter activity. Gprk2 RNAi flies are susceptible to infection with Enterococcus faecalis and Gprk2 RNAi rescues Toll(10b)-induced blood cell activation in Drosophila larvae in vivo. We conclude that Gprk2/GRK5 has an evolutionarily conserved role in regulating NF-kappaB signaling.
authors.affiliation: Institute of Medical Technology, University of Tampere, Tampere, Finland.
authors.forename: Susanna
authors.initials: S
authors.lastname: Valanne
authors.forename: Henna
authors.initials: H
authors.lastname: Myllymäki
authors.forename: Jenni
authors.initials: J
authors.lastname: Kallio
authors.forename: Martin Rudolf
authors.initials: MR
authors.lastname: Schmid
authors.forename: Anni
authors.initials: A
authors.lastname: Kleino
authors.forename: Astrid
authors.initials: A
authors.lastname: Murumägi
authors.forename: Laura
authors.initials: L
authors.lastname: Airaksinen
authors.forename: Tapio
authors.initials: T
authors.lastname: Kotipelto
authors.forename: Meri
authors.initials: M
authors.lastname: Kaustio
authors.forename: Johanna
authors.initials: J
authors.lastname: Ulvila
authors.forename: Shiva Seyedoleslami
authors.initials: SS
authors.lastname: Esfahani
authors.forename: Ylva
authors.initials: Y
authors.lastname: Engström
authors.forename: Olli
authors.initials: O
authors.lastname: Silvennoinen
authors.forename: Dan
authors.initials: D
authors.lastname: Hultmark
authors.forename: Mataleena
authors.initials: M
authors.lastname: Parikka
authors.forename: Mika
authors.initials: M
authors.lastname: Rämet
mesh_terms: D000818:Animals; D015153:Blotting, Western; D004330:Drosophila; D029721:Drosophila Proteins / Q000276:immunology* / Q000378:metabolism; D054769:G-Protein-Coupled Receptor Kinase 2 / Q000276:immunology* / Q000378:metabolism; D054774:G-Protein-Coupled Receptor Kinase 5 / Q000276:immunology / Q000378:metabolism*; D006091:Gram-Negative Chemolithotrophic Bacteria / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007113:Immunity, Innate; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D016328:NF-kappa B / Q000276:immunology* / Q000378:metabolism; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000502:physiology*; D015027:Zebrafish
chemical_list: D029721:Drosophila Proteins; D016328:NF-kappa B; D054769:G-Protein-Coupled Receptor Kinase 2; D054774:G-Protein-Coupled Receptor Kinase 5; C496574:Gprk2 protein, Drosophila