title: Intron cleavage affects processing of alternatively spliced transcripts.
abstract: We previously showed that the insertion of a hammerhead ribozyme (Rz) in a critical intronic position between the EDA exon and a downstream regulatory element affects alternative splicing. Here we evaluate the effect of other intronic cotranscriptional cleavage events on alternative pre-mRNA processing using different ribozymes (Rz) and Microprocessor target sequences (MTSs). In the context of the fibronectin EDA minigene, intronic MTSs were cleaved very inefficiently and did not affect alternative splicing or the level of mature transcripts. On the contrary, all hammerhead Rz derivatives and hepatitis δ Rz were completely cleaved before a splicing decision and able to affect alternative splicing. Despite the very efficient Rz-mediated cleavage, the levels of mature mRNA were only reduced to ∼40%. We show that this effect on mature transcripts occurs regardless of the type and intronic position of Rzs, or changes in alternative splicing and exon definition. Thus, we suggest that intron integrity is not strictly required for splicing but is necessary for efficient pre-mRNA biosynthesis.
authors.affiliation: International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34149 Trieste, Italy.
authors.forename: Tibor
authors.initials: T
authors.lastname: Pastor
authors.forename: Andrea
authors.initials: A
authors.lastname: Dal Mas
authors.forename: Gabriele
authors.initials: G
authors.lastname: Talotti
authors.forename: Erica
authors.initials: E
authors.lastname: Bussani
authors.forename: Franco
authors.initials: F
authors.lastname: Pagani
mesh_terms: D017398:Alternative Splicing; D045744:Cell Line, Tumor; D005786:Gene Expression Regulation; D006801:Humans; D007438:Introns; D016337:RNA, Catalytic / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D050436:Regulatory Elements, Transcriptional; D014158:Transcription, Genetic
chemical_list: D016337:RNA, Catalytic; D012333:RNA, Messenger; C427204:hammerhead ribozyme