Characterization of the human immunoglobulin epsilon mRNAs and their polyadenylation sites.

Abstract

Several IgE heavy (H) chain transcripts are produced by alternative splicing between constant region (CH3 and CH4) and membrane (M1 and M2) exons and by differential cleavage-polyadenylation at poly(A) sites downstream of the CH4 and M2 exons. We have now characterized the poly(A) signal of the epsilon transcripts that contain membrane exon sequences (epsilon CH4-M1'-M2, epsilon CH4-M1-M2, epsilon CH4-M2' and epsilon CH4-M2") and have determined the complete sequence of the M2 exon and 1.4 kb of downstream genomic DNA. The membrane locus poly(A) site was identified by RACE-PCR analysis of epsilon transcripts obtained from IgE-producing myeloma cells and normal peripheral blood lymphocytes (PBL). All membrane exon transcripts were found to be polyadenylated following a CA dinucleotide located 1046 nt from the beginning of the M2 exon. An AGTAAA hexamer, located 13 nt upstream from the site of cleavage and polyadenylation, was the only poly(A) signal sequence present in the 1.4 kb of genomic DNA downstream of the M2 exon. A (G+T)-rich region, which is also conserved in most poly(A) signals, was present 50 nt downstream of the AGTAAA hexamer. Northern blot analysis confirmed that this poly(A) site is used by the membrane exon epsilon mRNAs expressed by the U266 myeloma. The four membrane exon transcripts were detected in different relative amounts in PBL and IgE-producing myeloma cells, which could reflect different epsilon mRNA splicing patterns during B-cell differentiation.