Cloning of a novel phosphotyrosine binding domain containing molecule, Odin, involved in signaling by receptor tyrosine kinases.
We have used a proteomic approach using mass spectrometry to identify signaling molecules involved in receptor tyrosine kinase signaling pathways. Using affinity purification by anti-phosphotyrosine antibodies to enrich for tyrosine phosphorylated proteins, we have identified a novel signaling molecule in the epidermal growth factor receptor signaling pathway. This molecule, designated Odin, contains several ankyrin repeats, two sterile alpha motifs and a phosphotyrosine binding domain and is ubiquitously expressed. Using antibodies against endogenous Odin, we show that it undergoes tyrosine phosphorylation upon addition of growth factors such as EGF or PDGF but not by cytokines such as IL-3 or erythropoietin. Immunofluorescence experiments as well as Western blot analysis on subcellular fractions demonstrated that Odin is localized to the cytoplasm both before and after growth factor treatment. Deletion analysis showed that the phosphotyrosine binding domain of Odin is not required for its tyrosine phosphorylation. Overexpression of Odin, but not an unrelated adapter protein, Grb2, inhibited EGF-induced activation of c-Fos promoter. Microinjection of wild-type or a mutant version lacking the PTB domain into NIH3T3 fibroblasts inhibited PDGF-induced mitogenesis. Taken together, our results indicate that Odin may play a negative role in growth factor receptor signaling pathways.
Pandey
Akhilesh
A
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA. Pandey@cebi.sdu.dk
Blagoev
Blagoy
B
Kratchmarova
Irina
I
Fernandez
Minerva
M
Nielsen
Mogens
M
Kristiansen
Troels Zakarias
TZ
Ohara
Osamu
O
Podtelejnikov
Alexandre V
AV
Roche
Serge
S
Lodish
Harvey F
HF
Mann
Matthias
M
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000378:metabolism; D008969:Molecular Sequence Data; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D016760:Proto-Oncogene Proteins c-fos / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
C557872:ANKS1A protein, human; D048868:Adaptor Proteins, Signal Transducing; D010982:Platelet-Derived Growth Factor; D016760:Proto-Oncogene Proteins c-fos; D019000:Phosphotyrosine; D004815:Epidermal Growth Factor; D020794:Receptor Protein-Tyrosine Kinases