Shc contains two Grb2 binding sites needed for efficient formation of complexes with SOS in B lymphocytes.
Cross-linking of the B-cell antigen receptor (BCR) induces tyrosine phosphorylation of Shc, which is believed to lead to the activation of Ras. Previous work has shown that tyrosine-phosphorylated Shc forms complexes with another adapter protein, Grb2, and the Ras guanine nucleotide exchange factor SOS. Here, we demonstrate that phosphorylation of Shc by the hematopoietic cell-specific tyrosine kinase Syk induces binding of Grb2 to Shc, suggesting that Syk phosphorylates Shc in stimulated B cells. Surprisingly, Syk-phosphorylated Shc possesses two Grb2 binding sites rather than the one site that has been previously reported. Both of these sites are required for efficient formation of Shc-Grb2-SOS complexes in vitro and in vivo. We suggest that two Grb2 proteins anchored by a single Shc protein bind simultaneously to one SOS molecule, resulting in a complex that is more stable than a complex containing only a single Grb2 protein bound to one SOS molecule. This model is consistent with our observation that BCR stimulation greatly increases the amount of SOS associated with Grb2.
Harmer
S L
SL
G.W. Hooper Foundation and Department of Biochemistry and Biophysics, University of California, San Francisco 94143-0552, USA.
DeFranco
A L
AL
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000502:physiology*; D001665:Binding Sites; D002478:Cells, Cultured; D004792:Enzyme Precursors / Q000502:physiology; D051380:GRB2 Adaptor Protein; D019204:GTP-Binding Proteins / Q000502:physiology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000502:physiology*; D051379:Mice; D019000:Phosphotyrosine / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D011506:Proteins / Q000502:physiology*; D011940:Receptor Aggregation; D011947:Receptors, Antigen, B-Cell / Q000502:physiology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction; D020664:Son of Sevenless Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D000072377:Syk Kinase
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D004792:Enzyme Precursors; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; C495949:Grb2 protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D008565:Membrane Proteins; D011506:Proteins; D011947:Receptors, Antigen, B-Cell; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D020664:Son of Sevenless Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; C000606897:Syk protein, mouse; D019204:GTP-Binding Proteins