Abstract
Adaptor protein c-Abl SH3 domain-binding protein-2 (3BP2, also referred to SH3BP2) regulates immune receptor-mediated signal transduction. In this report we focused on the molecular mechanism of 3BP2 function in B cell receptor (BCR) signaling. Engagement of BCR induces tyrosine phosphorylation of 3BP2. Genetic analysis demonstrated that Syk is critical for BCR-mediated tyrosine phosphorylation of 3BP2. Mutational analysis of 3BP2 revealed that both Tyr(183) and Src homology 2 (SH2) domain are necessary for 3BP2-mediated BCR-induced activation of nuclear factor of activated T cells (NFAT). Point mutation of Tyr(183) or Arg(486) in the SH2 domain of 3BP2 diminished BCR-mediated tyrosine phosphorylation of 3BP2. Endogenous 3BP2 forms a complex with tyrosine-phosphorylated cellular signaling molecules. Peptide binding experiments demonstrated that only phosphorylated Tyr(183) in 3BP2 could form a complex with the SH2 domain(s) of phospholipase Cgamma2 and Vav1 from B cell lysates. These interactions were represented by using bacterial glutathione S-transferase-phospholipase Cgamma2 or -Vav1 SH2 domain. Furthermore, pulldown and Far Western experiments showed that the 3BP2-SH2 domain directly binds to B cell linker protein (BLNK) after BCR stimulation. These results demonstrated that 3BP2 induces the protein complex with cellular signaling molecules through phosphorylation of Tyr(183) and SH2 domain leading to the activation of NFAT in B cells.
Highlights
Ylation of cellular signaling molecules including B cell linker protein (BLNK), which forms a complex with Btk [1,2,3]
We previously demonstrated that adaptor protein 3BP2 is tyrosine-phosphorylated by a non-receptor type of protein-tyrosine kinase (PTK), Syk, Lyn, or Btk, which are known to be activated after B cell receptor (BCR) stimulation [30]
These findings suggested that tyrosine phosphorylation of 3BP2 may occur in the early events in BCR signaling pathway
Summary
Ylation of cellular signaling molecules including BLNK ( known as BASH or SLP-65), which forms a complex with Btk [1,2,3]. Both phosphorylation of Tyr183 and SH2 domain are required for BCR-mediated tyrosine phosphorylation of 3BP2 and activation of NFAT by associating specific signaling molecules. We previously demonstrated that adaptor protein 3BP2 is tyrosine-phosphorylated by a non-receptor type of PTKs, Syk, Lyn, or Btk, which are known to be activated after BCR stimulation [30].
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