Tyrosine phosphorylation of Sprouty2 enhances its interaction with c-Cbl and is crucial for its function.
Mammalian Sprouty (Spry) proteins are now established as receptor tyrosine kinase-induced modulators of the Ras/mitogen-activated protein kinase pathway. Specifically, hSpry2 inhibits the fibroblast growth factor receptor (FGFR)-induced mitogen-activated protein kinase pathway but conversely prolongs activity of the same pathway following epidermal growth factor (EGF) stimulation, where activated EGF receptors are retained on the cell surface. In this study it is demonstrated that hSpry2 is tyrosine-phosphorylated upon stimulation by either FGFR or EGF and subsequently binds endogenous c-Cbl with high affinity. A conserved motif on hSpry2, together with phosphorylation on tyrosine 55, is required for its enhanced interaction with the SH2-like domain of c-Cbl. A hSpry2 mutant (Y55F) that did not exhibit an enhanced binding with c-Cbl failed to retain EGF receptors on the cell surface. Furthermore, individually mutating hSpry2 residues 52-59 to alanine indicated a tight correlation between their affinity for c-Cbl binding and their inhibition of ERK2 activity in the FGFR pathway. We postulate that tyrosine phosphorylation "activates" hSpry2 by enhancing its interaction with c-Cbl and that this interaction is critical for its physiological function in a signal-specific context.
Fong
Chee Wai
CW
Signal Transduction Laboratory, Institute of Molecular and Cell Biology, National University of Singapore, Singapore, 117609 Singapore.
Leong
Hwei Fen
HF
Wong
Esther Sook Miin
ES
Lim
Jormay
J
Yusoff
Permeen
P
Guy
Graeme R
GR
D000409:Alanine / Q000737:chemistry; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D015536:Down-Regulation; D006801:Humans; D020935:MAP Kinase Signaling System; D018613:Microscopy, Confocal; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism* / Q000502:physiology; D050721:Proto-Oncogene Proteins c-cbl; D017468:Receptors, Fibroblast Growth Factor / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D018909:src Homology Domains
D018076:DNA, Complementary; D009419:Nerve Tissue Proteins; D011518:Proto-Oncogene Proteins; D017468:Receptors, Fibroblast Growth Factor; C404992:Spry2 protein, rat; D014443:Tyrosine; D004247:DNA; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D020928:Mitogen-Activated Protein Kinases; C067419:CBL protein, human; D000409:Alanine