Relationship between 3' end formation and SL2-specific trans-splicing in polycistronic Caenorhabditis elegans pre-mRNA processing.

About 25% of the genes in the nematode Caenorhabditis elegans are in operons, polycistronic transcription units in which the genes are only 100-400 bp apart. The operon pre-mRNAs are processed into monocistronic mRNAs by a combination of cleavage and polyadenylation at the 3' end of the upstream mRNA and SL2 trans-splicing at the 5' end of the downstream mRNA. To determine whether 3' end formation and SL2 trans-splicing are coupled mechanistically, we tested a gpd-2/gpd-3 operon construct driven by a C. elegans heat shock promoter, and measured the effects of inhibition of 3' end formation and/or trans-splicing on the processing of the polycistronic RNA in vivo. The results indicate that proper 3' end formation of the upstream mRNA in an operon is required for SL2-specificity of downstream mRNA trans-splicing. In contrast, trans-splicing of the downstream mRNA is not necessary for correct 3' end formation of the upstream mRNA. In addition, shortening the distance between the 5' cap and the AAUAAA of gpd-2 (the upstream gene) decreases the efficiency of 3' end formation and is accompanied by a replacement of SL2 with SL1 at the trans-splice site of gpd-3, the downstream gene. These results indicate that SL2 trans-splicing, in C. elegans, is coupled mechanistically to 3' end formation in the processing of polycistronic pre-mRNAs.