title: Differential protein occupancy profiling of the mRNA transcriptome.
abstract: BACKGROUND
  RNA-binding proteins (RBPs) mediate mRNA biogenesis, translation and decay. We recently developed an approach to profile transcriptome-wide RBP contacts on polyadenylated transcripts by next-generation sequencing. A comparison of such profiles from different biological conditions has the power to unravel dynamic changes in protein-contacted cis-regulatory mRNA regions without a priori knowledge of the regulatory protein component.
  
  
  RESULTS
  We compared protein occupancy profiles of polyadenylated transcripts in MCF7 and HEK293 cells. Briefly, we developed a bioinformatics workflow to identify differential crosslinking sites in cDNA reads of 4-thiouridine crosslinked polyadenylated RNA samples. We identified 30,000 differential crosslinking sites between MCF7 and HEK293 cells at an estimated false discovery rate of 10%. 73% of all reported differential protein-RNA contact sites cannot be explained by local changes in exon usage as indicated by complementary RNA-seq data. The majority of differentially crosslinked positions are located in 3' UTRs, show distinct secondary-structure characteristics and overlap with binding sites of known RBPs, such as ELAVL1. Importantly, mRNA transcripts with the most significant occupancy changes show elongated mRNA half-lives in MCF7 cells.
  
  
  CONCLUSIONS
  We present a global comparison of protein occupancy profiles from different cell types, and provide evidence for altered mRNA metabolism as a result of differential protein-RNA contacts. Additionally, we introduce POPPI, a bioinformatics workflow for the analysis of protein occupancy profiling experiments. Our work demonstrates the value of protein occupancy profiling for assessing cis-regulatory RNA sequence space and its dynamics in growth, development and disease.
authors.forename: Markus
authors.initials: M
authors.lastname: Schueler
authors.forename: Mathias
authors.initials: M
authors.lastname: Munschauer
authors.forename: Lea Haarup
authors.initials: LH
authors.lastname: Gregersen
authors.forename: Ana
authors.initials: A
authors.lastname: Finzel
authors.forename: Alexander
authors.initials: A
authors.lastname: Loewer
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.forename: Markus
authors.initials: M
authors.lastname: Landthaler
authors.forename: Christoph
authors.initials: C
authors.lastname: Dieterich
mesh_terms: D019295:Computational Biology; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D061986:MCF-7 Cells; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D017423:Sequence Analysis, RNA; D014158:Transcription, Genetic; D059467:Transcriptome
chemical_list: D012333:RNA, Messenger; D016601:RNA-Binding Proteins