The SH2/SH3 adaptor Grb4 transduces B-ephrin reverse signals.
Bidirectional signals mediated by membrane-anchored ephrins and Eph receptor tyrosine kinases have important functions in cell-cell recognition events, including those that occur during axon pathfinding and hindbrain segmentation. The reverse signal that is transduced into B-ephrin-expressing cells is thought to involve tyrosine phosphorylation of the signal's short, conserved carboxy-terminal cytoplasmic domain. The Src-homology-2 (SH2) domain proteins that associate with activated tyrosine-phosphorylated B-subclass ephrins have not been identified, nor has a defined cellular response to reverse signals been described. Here we show that the SH2/SH3 domain adaptor protein Grb4 binds to the cytoplasmic domain of B ephrins in a phosphotyrosine-dependent manner. In response to B-ephrin reverse signalling, cells increase FAK catalytic activity, redistribute paxillin, lose focal adhesions, round up, and disassemble F-actin-containing stress fibres. These cellular responses can be blocked in a dominant-negative fashion by expression of the isolated Grb4 SH2 domain. The Grb4 SH3 domains bind a unique set of other proteins that are implicated in cytoskeletal regulation, including the Cbl-associated protein (CAP/ponsin), the Abl-interacting protein-1 (Abi-1), dynamin, PAK1, hnRNPK and axin. These data provide a biochemical pathway whereby cytoskeletal regulators are recruited to Eph-ephrin bidirectional signalling complexes.
Cowan
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Center for Developmental Biology and Kent Waldrep Foundation Center for Basic Research on Nerve Growth and Regeneration, University of Texas Southwestern Medical Center, Dallas 75390-9133, USA.
Henkemeyer
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D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D060466:Axin Protein; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D036387:Ephrin-B1; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism; D015513:Oncogene Proteins / Q000737:chemistry / Q000502:physiology*; D019000:Phosphotyrosine / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D036224:Receptor, EphB4; D036081:Receptors, Eph Family; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins; D015398:Signal Transduction; D020798:Two-Hybrid System Techniques; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D060466:Axin Protein; D003598:Cytoskeletal Proteins; D036387:Ephrin-B1; D008565:Membrane Proteins; D008840:Microfilament Proteins; C485544:NCK2 protein, human; C117758:Nck2 protein, mouse; D015513:Oncogene Proteins; D011506:Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C119201:ponsin; D019000:Phosphotyrosine; D005982:Glutathione Transferase; D020794:Receptor Protein-Tyrosine Kinases; D036224:Receptor, EphB4; D036081:Receptors, Eph Family