ABSTRACT
B lymphocyte-restricted transcription of immunoglobulin heavy-chain (IgH) genes is specified by elements within the variable region (VH) promoter and the intronic enhancer (E mu). The gene encoding a protein that binds a VH promoter proximal site necessary for induced mu-heavy-chain transcription has been cloned. This B-cell specific protein, termed Bright (B cell regulator of IgH transcription), is found in both soluble and matrix insoluble nuclear fractions. Bright binds the minor groove of a restricted ATC sequence that is sufficient for nuclear matrix association. This sequence motif is present in previously described matrix-associating regions (MARs) proximal to the promoter and flanking E mu. Bright can activate E mu-driven transcription by binding these sites, but only when they occur in their natural context and in cell lines permissive for E mu activity. To bind DNA, Bright requires a novel tetramerization domain and a previously undescribed domain that shares identity with several proteins, including SWI1, a component of the SWI/SNF complex.
Subject(s)
B-Lymphocytes/metabolism , DNA-Binding Proteins/metabolism , Immunoglobulin Heavy Chains/metabolism , Oncogenes , Trans-Activators/metabolism , Amino Acid Sequence , Base Sequence , Cell Line , Cells, Cultured , Cloning, Molecular , DNA, Complementary , DNA-Binding Proteins/genetics , Enhancer Elements, Genetic , Gene Expression Regulation, Developmental , Immunoglobulin Heavy Chains/genetics , Molecular Sequence Data , Protein Binding , Sequence Homology, Amino Acid , Trans-Activators/genetics , Transcription FactorsABSTRACT
Potentiation of initial signal transduction events through the cross-linking of the B cell antigen receptor complex appears to be dependent upon the association of membrane immunoglobulin (mIg) with Ig alpha and Ig beta. We made two groups of mutations within the COOH terminus of mIgM substituting: 1) the spacer, transmembrane, and cytoplasmic domains and 2) the NH2-terminal 2-8 amino acids within the transmembrane domain (NLWTTAST). We then evaluated the ability of the mutated receptors to associate with Ig alpha and Ig beta and to initiate signal transduction events (Ca2+ mobilization and phosphorylation by tyrosine protein kinases) after cross-linking mIgM receptors. Mutant mIgM receptors containing substitutions of gamma 2b (spacer, transmembrane, and cytoplasmic domains), AA for TT, and AAAAA for TTAST bound Ig alpha and Ig beta and initiated signal transduction events after mIgM receptor cross-linking. However, substitutions of I-A alpha (spacer, transmembrane, and cytoplasmic domains) or TTVVCALGL for NLWTTAST blocked association of Ig alpha and Ig beta and initiation of signal transduction events. Results indicate that residues within the first 8 amino acids of the transmembrane domain other than TTAST are necessary for receptor function and association with Ig alpha and Ig beta.
