Differential protein occupancy profiling of the mRNA transcriptome
Research output: Contribution to journal › Journal article › Research › peer-review
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.
Original language | English |
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Article number | R15 |
Journal | Genome Biology |
Volume | 15 |
Issue number | 1 |
ISSN | 1474-7596 |
DOIs | |
Publication status | Published - 13 Jan 2014 |
Externally published | Yes |
- Computational Biology, Gene Expression Regulation, HEK293 Cells, High-Throughput Nucleotide Sequencing, Humans, MCF-7 Cells, RNA, Messenger/genetics, RNA-Binding Proteins/genetics, Sequence Analysis, RNA, Transcription, Genetic, Transcriptome
Research areas
ID: 328236641