PUBLICATION

Genome-Wide RNA Interference in Drosophila Cells Identifies G Protein-Coupled Receptor Kinase 2 as a Conserved Regulator of NF-{kappa}B Signaling

Authors
Valanne, S., Myllymäki, H., Kallio, J., Schmid, M.R., Kleino, A., Murumägi, A., Airaksinen, L., Kotipelto, T., Kaustio, M., Ulvila, J., Esfahani, S.S., Engström, Y., Silvennoinen, O., Hultmark, D., Parikka, M., and Rämet, M.
ID
ZDB-PUB-100504-4
Date
2010
Source
Journal of immunology (Baltimore, Md. : 1950)   184(11): 6188-6198 (Journal)
Registered Authors
Keywords
none
MeSH Terms
  • Humans
  • Zebrafish
  • G-Protein-Coupled Receptor Kinase 2/immunology*
  • G-Protein-Coupled Receptor Kinase 2/metabolism
  • NF-kappa B/immunology*
  • NF-kappa B/metabolism
  • Drosophila
  • Drosophila Proteins/immunology*
  • Drosophila Proteins/metabolism
  • Gram-Negative Chemolithotrophic Bacteria/immunology
  • Gram-Negative Chemolithotrophic Bacteria/metabolism
  • Blotting, Western
  • Immunity, Innate*
  • RNA Interference
  • G-Protein-Coupled Receptor Kinase 5/immunology
  • G-Protein-Coupled Receptor Kinase 5/metabolism*
  • Immunohistochemistry
  • Immunoprecipitation
  • Reverse Transcriptase Polymerase Chain Reaction
  • Animals
  • Signal Transduction/physiology*
(all 21)
PubMed
20421637 Full text @ 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.
Genes / Markers
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Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Antibodies
Orthology
Engineered Foreign Genes
Mapping
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Citation
Valanne, S., Myllymäki, H., Kallio, J., Schmid, M.R., Kleino, A., Murumägi, A., Airaksinen, L., Kotipelto, T., Kaustio, M., Ulvila, J., Esfahani, S.S., Engström, Y., Silvennoinen, O., Hultmark, D., Parikka, M., and Rämet, M. (2010) Genome-Wide RNA Interference in Drosophila Cells Identifies G Protein-Coupled Receptor Kinase 2 as a Conserved Regulator of NF-{kappa}B Signaling. Journal of immunology (Baltimore, Md. : 1950). 184(11):6188-6198.