RRC ID 11070
Author Valanne S, Myllymäki H, Kallio J, Schmid MR, Kleino A, Murumägi A, Airaksinen L, Kotipelto T, Kaustio M, Ulvila J, Esfahani SS, Engström Y, Silvennoinen O, Hultmark D, Parikka M, Rämet M.
Title Genome-wide RNA interference in Drosophila cells identifies G protein-coupled receptor kinase 2 as a conserved regulator of NF-kappaB signaling.
Journal J. Immunol.
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.
Volume 184(11)
Pages 6188-98
Published 2010-6-1
DOI 10.4049/jimmunol.1000261
PII jimmunol.1000261
PMID 20421637
MeSH Animals Blotting, Western Drosophila Drosophila Proteins / immunology* Drosophila Proteins / metabolism G-Protein-Coupled Receptor Kinase 2 / immunology* G-Protein-Coupled Receptor Kinase 2 / metabolism G-Protein-Coupled Receptor Kinase 5 / immunology G-Protein-Coupled Receptor Kinase 5 / metabolism* Gram-Negative Chemolithotrophic Bacteria / immunology Gram-Negative Chemolithotrophic Bacteria / metabolism Humans Immunity, Innate* Immunohistochemistry Immunoprecipitation NF-kappa B / immunology* NF-kappa B / metabolism RNA Interference Reverse Transcriptase Polymerase Chain Reaction Signal Transduction / physiology* Zebrafish
IF 4.539
Times Cited 46
Drosophila 17998R-1 17998R-3