RESUMEN
Proteins that bind IgE play important roles in both the synthesis and function of IgE are therefore intimately involved in IgE-mediated human allergic disorders. This report describes the structure of an IgE-binding protein, as predicted from sequencing a cDNA cloned from rat basophilic leukemia cells. This protein contains two domains: the amino-terminal domain (140 amino acids) consists of a highly conserved repetitive amino acid sequence, Tyr-Pro-Gly-Pro/Gln-Ala/Thr-Pro/Ala-Pro-Gly-Ala, whereas the carboxyl-terminal domain (122 amino acids) shares significant sequence homology with a domain of lymphocyte/macrophage receptor for the Fc portion of IgG. Other proteins with this type of structure but with affinity for other immunoglobulin isotypes may exist and may represent a heretofore unidentified component of the immune system.
Asunto(s)
Inmunoglobulina E/inmunología , Inmunoglobulina G/inmunología , Receptores Fc/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Línea Celular , Datos de Secuencia Molecular , Receptores de IgE , Receptores de IgG , Secuencias Repetitivas de Ácidos NucleicosRESUMEN
The role of immunoglobulin structural genes in the generation of autoantibodies in humans has not been elucidated. Human monoclonal IgM anti-IgG autoantibodies (rheumatoid factors, RFs) from unrelated people often share idiotypic antigens. Antibodies against synthetic peptides have localized two of the shared idiotypic determinants to the second and third complementarity-determining regions of the kappa light chain. The reported sequences of several human RF light chains are remarkably homologous in these regions. Animal studies have shown that some shared idiotypic antigens represent serological markers for immunoglobulin variable (V)-region genes. Therefore, we hypothesized that human RF light chains derived from a single germ-line gene, designated V kappa-(RF), or from a small family of very closely related genes. In the present experiments, we have isolated and sequenced two human V kappa germ-line genes that encode kappa light chains, which are identical or closely related to the light chains of human RF. The data indicate that the shared idiotypic antigens on RF are phenotypic markers for a kappa V-region gene that is highly conserved in the human population. The results also imply that the light chains of IgM anti-IgG autoantibodies can be encoded by germ-line genes without any somatic mutation.
Asunto(s)
Idiotipos de Inmunoglobulinas/genética , Región Variable de Inmunoglobulina/genética , Cadenas kappa de Inmunoglobulina/genética , Factor Reumatoide/genética , Secuencia de Aminoácidos , Secuencia de Bases , Reacciones Cruzadas , ADN/aislamiento & purificación , Humanos , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunologíaRESUMEN
The synthesis and function of IgE are dependent on IgE-binding proteins, which include cell surface IgE receptors and IgE-binding lymphokines. To further our understanding of the IgE system, we have engaged in the molecular cloning of genes for some of these proteins. In studying the in vitro translation products of mRNA from rat basophilic leukemia (RBL) cells, we have identified a Mr 31,000 polypeptide that binds IgE and is also reactive with antibodies to proteins affinity-purified from RBL cells with IgE immunoadsorbent. For the molecular cloning, double-stranded cDNA 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, we identified one clone containing cDNA that hybridized to mRNA coding for a Mr 31,000 IgE-binding protein. 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 Mr 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.