CHARACTERIZATION OF THE HUMAN-IMMUNOGLOBULIN-EPSILON MESSENGER-RNAS AND THEIR POLYADENYLATION SITES

Several IgE heavy (H) chain transcripts are produced by alternative sp licing between constant region (CH3 and CH4) and membrane (M1 and M2) exons and by differential cleavage-polyadenylation at poly(A) sites do wnstream of the CH4 and M2 exons. We have now characterized the poly(A ) signal of the epsilon transcripts that contain membrane exon sequenc es (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 a nd 1.4 kb of downstream genomic DNA. The membrane locus poly(A) site w as identified by RACE-PCR analysis of epsilon transcripts obtained fro m IgE-producing myeloma cells and normal peripheral blood lymphocytes (PBL). All membrane exon transcripts were found to be polyadenylated f ollowing a CA dinucleotide located 1046 nt from the beginning of the M 2 exon. An AGTAAA hexamer, located 13 nt upstream from the site of cle avage and polyadenylation, was the only poly(A) signal sequence presen t 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 confirm ed that this poly(A) site is used by the membrane exon E mRNAs express ed by the U266 myeloma. The four membrane exon transcripts were detect ed in different relative amounts in PBL and IgE-producing myeloma cell s, which could reflect different E mRNA splicing patterns during B-cel l differentiation.