Spliceosomal small nuclear ribonucleoproteins in trypansomes: Genomwide analysis of role of U1 snRPN proteins in mRNA processing

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All protein-coding genes in trypanosomes are expressed as polycistronic primary transcripts that have to be processed by coupled trans splicing and polyadenylation to yield mature mRNAs. In addition, only very few genes are known to undergo cis splicing of an internal intron. Trans splicing requires specifically the Spliced Leader (SL) RNA, which adds its 5’-terminal 39-nucleotide miniexon to each of the protein-coding exons. On the other hand, the U1 small nuclear ribonucleoprotein (snRNP) is considered a component acting specifically during cis splicing of the very few internal introns. We have recently characterized the U1 snRNP from Trypanosoma brucei, an unusual RNP with a minimal U1 snRNA, U1-70K, U1C, and a protein component (U1-24K) not found in well-characterized U1 snRNPs from other systems. Evidence from several studies including our own has accumulated that argues for the existence of several different RNP complexes of the U1 snRNA and for U1-independent roles of their protein components, linking cis and trans splicing as well as polyadenylation. We propose to study in detail the contributions of individual U1 snRNP proteins to and the potential links between these principal mRNA processing reactions in trypanosomes, using systematic epitope tagging, RNAi knockdown, as well as genomewide RNA-binding (iCLIP) and deep-sequencing approaches.