B-Cell Activation by Wild Type and Mutant Ig-β Cytoplasmic Domains
In B lymphocytes, the cytoplasmic domains of the membrane immunoglobulin-associated heterodimeric Ig-α and Ig-β proteins link membrane immunoglobulin to intracellular signalling molecules. We constructed chimeric genes encoding the extracellular and transmembrane domain of human CD8α and the cytoplasmic domain of Ig-α or Ig-β and examined the ability of the chimeric proteins to induce signalling in the murine B-cell lymphoma A20. Crosslinking of CD8/Ig-α or CD8/Ig-β induced both calcium mobilization and protein tyrosine phosphorylation, although induction by CD8/Ig-α was somewhat stronger. We also carried out mutagenesis of residues within the “Reth” motif of the CD8/Ig-β cytoplasmic domain and determined the effects of these mutations on signalling in the murine B-cell hybridoma LK 35.2. Mutants in which alanine was substituted for glutamine 202, threonine 205, and isoleucine 209 retained the ability to induce protein tyrosine phosphorylation and calcium mobilization. In contrast, substitution of alanine for leucine 198 abrogated these responses, suggesting a critical role for this residue in interaction with cytoplasmic signalling proteins.
KeywordsCytoplasmic Domain Antigen Receptor Chimeric Protein Vesicular Stomatitis Virus Bovine Leukemia Virus
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