Characterization and expression of alternatively spliced IgE heavy chain transcripts produced by peripheral blood lymphocytes.
We have investigated the IgE heavy chain isoforms produced in vivo by analyzing the epsilon mRNA species present in unstimulated PBL and expressing them individually in a myeloma cell line. Seven epsilon mRNA species were identified by using reverse transcription-PCR, cloning, and sequencing analysis. These species included the classical secreted (epsilon CH4-S) and membrane-bound (epsilon CH4-M1'-M2) IgE and five alternatively spliced epsilon transcripts. At the protein level, the five alternatively spliced epsilon transcripts (epsilon CH4*, epsilon CH4-M2', epsilon CH4'-1, epsilon CH4'-1-M2, and epsilon CH3-13-CH4) corresponded to four epsilon heavy chain isoforms, in which various parts of the CH4 domain were replaced by new stretches of amino acids at the carboxyl termini. The same epsilon mRNA species also were present in the IgE producing myeloma cell line U266. However, except for the classical membrane and secreted IgE, the corresponding proteins could not be identified. To further characterize the epsilon CH4-S, epsilon CH4*, epsilon CH4-M2', epsilon CH4'-1, and epsilon CH4-M1'-M2 species, we expressed them as chimeric mouse/human anti-4-hydroxy-5-iodo-3-nitrophenacetyl Abs in a mouse myeloma cell line. Only the classical secreted and membrane isoforms were found to be secreted or expressed on the cell surface, respectively, and the other forms were retained within the cells and degraded. These data suggest that some of the epsilon mRNA isoforms produced by PBL are aberrantly spliced mRNAs, the protein products of which are eliminated by post-translational events.
Batista
F D
FD
Molecular Immunology Group, International Center for Genetic Engineering and Biotechnology, Trieste, Italy.
Efremov
D G
DG
Burrone
O R
OR
D000595:Amino Acid Sequence; D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology*; D001483:Base Sequence; D003001:Cloning, Molecular; D005803:Genes, Immunoglobulin; D006801:Humans; D007073:Immunoglobulin E / Q000096:biosynthesis / Q000235:genetics*; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009101:Multiple Myeloma; D009194:Myeloma Proteins / Q000096:biosynthesis / Q000235:genetics; D010954:Plasmacytoma; D016133:Polymerase Chain Reaction; D012326:RNA Splicing; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014407:Tumor Cells, Cultured
D007143:Immunoglobulin Heavy Chains; D009194:Myeloma Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D007073:Immunoglobulin E