Formation of Shc/Grb2- and Crk adaptor complexes containing tyrosine phosphorylated Cbl upon stimulation of the B-cell antigen receptor.
B-cell antigen receptor (BCR) stimulation induces tyrosine phosphorylation of the Shc adaptor protein and its association with Grb2. The Shc/Grb2 complex may be involved in Ras activation, since Grb2 interacts with the guanine nucleotide exchange factor Sos. We reveal here an additional complexity of the BCR-induced Shc/Grb2 complex: it contains tyrosine phosphorylated proteins of 130, 110 and 75 kDa. The 130 kDa molecule inducibly associates with Shc, while the 75 kDa protein interacts with the carboxy-terminal SH3 domain of Grb2. The 110 kDa molecule is defined as Cbl, the product of the c-cbl oncogene, which is strongly phosphorylated on tyrosine upon BCR stimulation. Cbl constitutively interacts with the SH3 domains of Grb2, with a preference for the amino-terminal domain, and is in this way recruited to Shc upon BCR stimulation. Immunodepletion studies showed that Grb2-associated Cbl can be phosphorylated by BCR-induced tyrosine kinases independent of a Shc/Grb2 interaction. This indicates that the BCR can also couple to a Grb2 complex without the involvement of Shc. Cbl not only interacts with Grb2, but also with the adaptor protein Crk. In contrast to its constitutive interaction with Grb2, tyrosine-phosphorylated Cbl only associates with Crk after BCR stimulation. In summary, we observe that the BCR activates Shc/Grb2-, Grb2- and Crk adaptor complexes of distinct composition, which may allow selective coupling to different signal transduction cascades. Cbl participates in all three adaptor complexes and is tyrosine phosphorylated upon BCR stimulation, pointing to a central role for this molecule in the regulation of antigen receptor-induced B cell responses.
Smit
L
L
Division of Cellular Biochemistry, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
van der Horst
G
G
Borst
J
J
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000595:Amino Acid Sequence; D001483:Base Sequence; D002051:Burkitt Lymphoma / Q000378:metabolism; D029968:Escherichia coli Proteins; D051380:GRB2 Adaptor Protein; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010766:Phosphorylation; D014176:Protein Biosynthesis; D011506:Proteins / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000378:metabolism; D050737:Proto-Oncogene Proteins c-crk; D011947:Receptors, Antigen, B-Cell / Q000502:physiology*; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction / Q000502:physiology; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014157:Transcription Factors / Q000096:biosynthesis / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism*; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; C097229:Cbl protein, E coli; D029968:Escherichia coli Proteins; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D046911:Macromolecular Substances; D011506:Proteins; D011518:Proto-Oncogene Proteins; D050737:Proto-Oncogene Proteins c-crk; D011947:Receptors, Antigen, B-Cell; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014157:Transcription Factors; D014443:Tyrosine