title: The adaptor protein Bam32 regulates Rac1 activation and actin remodeling through a phosphorylation-dependent mechanism.
abstract: The B cell adaptor molecule of 32 kDa (Bam32) is an adaptor that links the B cell antigen receptor (BCR) to ERK and JNK activation and ultimately to mitogenesis. After BCR cross-linking, Bam32 is recruited to the plasma membrane and accumulates within F-actin-rich membrane ruffles. Bam32 contains one Src homology 2 and one pleckstrin homology domain and is phosphorylated at a single site, tyrosine 139. To define the function of Bam32 in membrane-proximal signaling events, we established human B cell lines overexpressing wild-type or mutant Bam32 proteins. The basal level of F-actin increased in cells expressing wild-type or myristoylated Bam32 but decreased in cells expressing either an Src homology-2 or Tyr-139 Bam32 mutant. Overexpression of wild-type Bam32 also affected BCR-induced actin remodeling, which was visualized as increases in F-actin-rich membrane ruffles. In contrast, Bam32 mutants largely blocked the BCR-induced increase in cellular F-actin. The positive and negative effects of Bam32 variants on F-actin levels were closely mirrored by their effects on the activation of the GTPase Rac1, which is known to regulate actin remodeling in lymphocytes. Bam32-deficient DT40 B cells showed decreased Rac1 activation and a failure of Rac1 to co-localize with the BCR, whereas cells overexpressing Bam32 had increased constitutive Rac1 activation. These results suggest that Bam32 regulates the cytoskeleton through Rac1. Bam32 variants also affected downstream signaling to JNK in a manner similar to that of Rac1, suggesting that the effect of Bam32 on JNK activation may be at least partially mediated through Rac1. Our results demonstrate a novel phosphorylation-dependent function of Bam32 in regulating Rac1 activation and actin remodeling.
authors.affiliation: Department of Immunology, University of Manitoba, Winnipeg, Manitoba R3E 0W3, Canada.
authors.forename: Atef
authors.initials: A
authors.lastname: Allam
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Niiro
authors.forename: Edward A
authors.initials: EA
authors.lastname: Clark
authors.forename: Aaron J
authors.initials: AJ
authors.lastname: Marshall
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D002051:Burkitt Lymphoma; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases; D008074:Lipoproteins / Q000235:genetics / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016296:Mutagenesis; D010766:Phosphorylation; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*; D018909:src Homology Domains / Q000235:genetics
chemical_list: D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D003432:Cross-Linking Reagents; C486555:DAPP1 protein, human; D008074:Lipoproteins; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D020830:rac1 GTP-Binding Protein