Regulation of process retraction and cell migration by EphA3 is mediated by the adaptor protein Nck1.
The Eph family of tyrosine kinase receptors and their ligands, the ephrins, participates in the regulation of a wide variety of biological functions under normal and pathological conditions. During embryonic development, interactions between the ligands and receptors define tissue boundaries, guide migrating axons, and regulate angiogenesis, as well as bone morphogenesis. These molecules have also been shown to modify neural activity in the adult nervous system and influence tumor progression. However, the molecular mechanisms underlying these diverse functions are not completely understood. In this study, we conducted a yeast two-hybrid screen to identify molecules that physically interact with Eph receptors using the cytoplasmic domain of EphA3 as "bait". This study identified Nck1 as a strong binding partner of EphA3 as assayed using both GST fusion protein pull down and co-immunoprecipitation techniques. The interaction is mediated through binding of the Nck1 SH2 domain to the phosphotyrosine residue at position 602 (Y602) of the EphA3 receptor. The removal of the SH2 domain or the mutation of the Y602 residue abolishes the interaction. We further demonstrated that EphA3 activation inhibits cell migration and process outgrowth, and these inhibiting effects are partially alleviated by dominant-negative Nck1 mutants that lack functional SH2 or SH3 domains, but not by the wild-type Nck1 gene. These results suggest that Nck1 interacts with EphA3 to regulate cell migration and process retraction.
Hu
Tianjing
T
Department of Chemical Biology, College of Pharmacy, Rutgers University, Piscataway, New Jersey 08854, USA.
Shi
Guanfang
G
Larose
Louise
L
Rivera
Gonzalo M
GM
Mayer
Bruce J
BJ
Zhou
Renping
R
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000502:physiology*; D002465:Cell Movement / Q000502:physiology*; D047468:Immunoprecipitation; D015513:Oncogene Proteins / Q000737:chemistry / Q000502:physiology*; D036121:Receptor, EphA3 / Q000502:physiology*; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; C078332:Nck protein; D015513:Oncogene Proteins; D036121:Receptor, EphA3