Structural and functional basis of a role for CRKL in a fibroblast growth factor 8-induced feed-forward loop.
The adapter protein CRKL is required for the normal development of multiple tissues that rely on fibroblast growth factor 8 (FGF8). The precise role of CRKL in receptor signaling has been unclear, however. To address this issue, we first modeled the three-dimensional structure of CRKL by molecular dynamics. By taking advantage of structural simulations, we performed in silico analysis of the interactions of the autophosphorylation sites of FGR receptor 1 (FGFR1) with the SH2 domain of CRKL or a highly related protein, CRK. As predicted by simulations, we confirm the specific physical interaction of phosphorylated Y463 (pY463) in FGFR1 with the CRKL SH2 domain at an affinity approximately 30-fold stronger than that of CRK. We also provide evidence that interactions outside of the core YXXP motif have a significant impact on physical association, which is consistent with predictions from molecular-dynamics simulations. Furthermore, we identify CRKL as an essential component of an FGF8-induced feed-forward loop permissive for efficient activation of the mitogen-activated protein kinase Erk1/2, as well as FGF8-induced anchorage-independent cell growth, using Crkl-deficient cells or a pY463 synthetic peptide. Although many cells generally require cell-matrix adhesion, our results demonstrate that CRKL permits cells to bypass the strict need for adhesion in response to FGF8 through direct interaction with receptor.
Seo
Ji-Heui
JH
Ben May Department for Cancer Research, University of Chicago, Chicago, Illinois 60637, USA.
Suenaga
Atsushi
A
Hatakeyama
Mariko
M
Taiji
Makoto
M
Imamoto
Akira
A
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D019295:Computational Biology; D003198:Computer Simulation; D025461:Feedback, Physiological / Q000187:drug effects*; D051524:Fibroblast Growth Factor 8 / Q000494:pharmacology*; D006801:Humans; D048369:MAP Kinase Kinase 1 / Q000378:metabolism; D051379:Mice; D008954:Models, Biological; D008958:Models, Molecular; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D050737:Proto-Oncogene Proteins c-crk / Q000737:chemistry / Q000378:metabolism; D051496:Receptor, Fibroblast Growth Factor, Type 1 / Q000378:metabolism; D013329:Structure-Activity Relationship; D048490:raf Kinases / Q000378:metabolism; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; C485285:CRK protein, human; C082869:CRKL protein; D009687:Nuclear Proteins; D010455:Peptides; D050737:Proto-Oncogene Proteins c-crk; D051524:Fibroblast Growth Factor 8; D019000:Phosphotyrosine; D051496:Receptor, Fibroblast Growth Factor, Type 1; D048490:raf Kinases; D048369:MAP Kinase Kinase 1; D020559:Monomeric GTP-Binding Proteins