Identification of an IgE-binding protein by molecular cloning.

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Abstract
The synthesis and function of IgE are dependent on IgE-binding proteins, which include cell surface IgE receptors and IgE-binding lymphokines. To further understanding of the IgE system, the molecular cloning of genes for some of these proteins was undertaken. In studying the in vitro translation products of mRNA from rat basophilic leukemia (RBL) cells, a MW 31,000 polypeptide that binds IgE and is also reactive with antibodies to proteins affinity-purified from RBL cells with IgE immunoadsorbent was identified. For the molecular cloning, double-stranded cDNA [complementary DNA] was synthesized from sucrose gradient-fractionated RBL mRNA, inserted into plasmid pBR322, and used to transform Escherichia coli. By screening transformants with a hybridization-selection/in vitro translation procedure, 1 clone containing cDNA that hybridized to mRNA coding for a MW 31,000 IgE-binding protein was identified. The DNA sequence of this cloned cDNA (571 base pairs) was determined and the amino acid sequence corresponding to part of the protein was deduced. In RNA blot analysis, the cDNA hybridized with a mRNA of 1100 nucleotides found in RBL cells but absent in cells not expressing IgE receptors. This cloned cDNA most likely codes for the MW 31,000 IgE-binding protein identified in RBL cells, which appears to be related to the IgE-binding phenotype of the cells and which may have a significant role in the IgE-mediated activation of basophils and mast cells.