Splicing of Balbiani ring 1 gene pre-mRNA occurs simultaneously with transcription

The 40 kb Balbiani ring 1 (BR1) gene is at a given moment transcribed by, on average, 120 RNA polymerases. Here we directly assay the excision of introns both in the nascent and in the released nucleoplasmic BR1 pre-mRNAs, isolated by microdissection. We show that intron 3, located 3 kb from the 5' end of the pre-mRNA, is excised simultaneous with transcription. Within 2.5 min of transcription time, 50% of the pre-mRNA molecules have lost the intron. Intron 4, located 600 bases from the polyadenylation site, is excised cotranscriptionally in 5%-10% of the molecules and after or during release to the nucleoplasm in the remaining molecules. Our results demonstrate that spliceosome assembly is a cotranscriptional process in vivo and that splicing may occur during transcription but also after completed transcription, depending on the position of the intron.