A secondary structure at the 3' splice site affects the in vitro splicing reaction of mouse immunoglobulin mu chain pre-mRNAs

Abstract

The expression of the IgM (immunoglobulin mu) heavy chain gene is known to be regulated at the post-transcriptional level. The two isoforms, the membrane-bound and secreted forms, are generated from the same gene by alternative processing at the 3' end of the primary transcript. The processing reactions involved are polyadenylation at the upstream poly(A) site (for the secreted form) and polyadenylation at the downstream poly(A) site coupled with splicing between exon C4 and exon M1 (for the membrane-bound form). The regulatory mechanism underlying these differential processing reactions is still not well understood. We investigated the splicing reaction between exon C4 and exon M1 in a HeLa nuclear extract using model transcripts containing the 5' and 3' splice sites of the C4-M1 intron. We found that the 3' splice site of the C4-M1 intron is sequestered in a stem-loop structure, which inhibits the splicing reaction in vitro. The inhibition by the stem-loop structure was also observed with a mouse lymphoma extract.