RESUMO
Maturation of eukaryotic pre-mRNAs via splicing and polyadenylation is modulated across cell types and conditions by a variety of RNA-binding proteins (RBPs). Although there exist over 1,500 RBPs in human cells, their binding motifs and functions still remain to be elucidated, especially in the complex environment of tissues and in the context of diseases. To overcome the lack of methods for the systematic and automated detection of sequence motif-guided pre-mRNA processing regulation from RNA sequencing (RNA-Seq) data we have developed MAPP (Motif Activity on Pre-mRNA Processing). Applying MAPP to RBP knock-down experiments reveals that many RBPs regulate both splicing and polyadenylation of nascent transcripts by acting on similar sequence motifs. MAPP not only infers these sequence motifs, but also unravels the position-dependent impact of the RBPs on pre-mRNA processing. Interestingly, all investigated RBPs that act on both splicing and 3' end processing exhibit a consistently repressive or activating effect on both processes, providing a first glimpse on the underlying mechanism. Applying MAPP to normal and malignant brain tissue samples unveils that the motifs bound by the PTBP1 and RBFOX RBPs coordinately drive the oncogenic splicing program active in glioblastomas demonstrating that MAPP paves the way for characterizing pre-mRNA processing regulators under physiological and pathological conditions.
Assuntos
Poliadenilação , Precursores de RNA , Splicing de RNA , Proteínas de Ligação a RNA , Humanos , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/genética , Precursores de RNA/metabolismo , Precursores de RNA/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias/genética , Neoplasias/metabolismo , Motivos de Nucleotídeos , Proteína de Ligação a Regiões Ricas em Polipirimidinas/metabolismo , Proteína de Ligação a Regiões Ricas em Polipirimidinas/genética , Fatores de Processamento de RNA/metabolismo , Fatores de Processamento de RNA/genética , Ribonucleoproteínas Nucleares Heterogêneas/metabolismo , Ribonucleoproteínas Nucleares Heterogêneas/genética , RNA Mensageiro/metabolismo , RNA Mensageiro/genéticaRESUMO
The usage of alternative terminal exons results in messenger RNA (mRNA) isoforms that differ in their 3' untranslated regions (3' UTRs) and often also in their protein-coding sequences. Alternative 3' UTRs contain different sets of cis-regulatory elements known to regulate mRNA stability, translation and localization, all of which are vital to cell identity and function. In previous work, we revealed that â¼25 percent of the experimentally observed RNA 3' ends are located within regions currently annotated as intronic, indicating that many 3' end isoforms remain to be uncovered. Also, the inclusion of not yet annotated terminal exons is more tissue specific compared to the already annotated ones. Here, we present the single cell-based Terminal Exon Annotation database (scTEA-db, www.scTEA-db.org) that provides the community with 12 063 so far not yet annotated terminal exons and associated transcript isoforms identified by analysing 53 069 publicly available single cell transcriptomes. Our scTEA-db web portal offers an array of features to find and explore novel terminal exons belonging to 5538 human genes, 110 of which are known cancer drivers. In summary, scTEA-db provides the foundation for studying the biological role of large numbers of so far not annotated terminal exon isoforms in cell identity and function.