Abstract
The past decades have witnessed a surge of discoveries revealing RNA regulation as a central player in cellular processes. RNAs are regulated by RNA-binding proteins (RBPs) at all post-transcriptional stages, including splicing, transportation, stabilization and translation. Defects in the functions of these RBPs underlie a broad spectrum of human pathologies. Systematic identification of RBP functional targets is among the key biomedical research questions and provides a new direction for drug discovery. The advent of cross-linking immunoprecipitation coupled with high-throughput sequencing (genome-wide CLIP) technology has recently enabled the investigation of genome-wide RBP–RNA binding at single base-pair resolution. This technology has evolved through the development of three distinct versions: HITS-CLIP, PAR-CLIP and iCLIP. Meanwhile, numerous bioinformatics pipelines for handling the genome-wide CLIP data have also been developed. In this review, we discuss the genome-wide CLIP technology and focus on bioinformatics analysis. Specifically, we compare the strengths and weaknesses, as well as the scopes, of various bioinformatics tools. To assist readers in choosing optimal procedures for their analysis, we also review experimental design and procedures that affect bioinformatics analyses.
Highlights
The diversity of RNA in sequence and structure underpins much of cell heterogeneity and complexity
RNA-binding proteins (RBPs) are proteins that bind to double- or singlestranded RNAs in cells and form ribonucleoprotein complexes with the bound RNAs
There are several related technologies, such as CLASH and RNA immunoprecipitation sequencing (RIP-Seq), which may be complementary to genome-wide CLIP to study the function of RNAs
Summary
The diversity of RNA in sequence and structure underpins much of cell heterogeneity and complexity. RNA-binding proteins (RBPs) are proteins that bind to double- or singlestranded RNAs in cells and form ribonucleoprotein complexes with the bound RNAs. Located in either the nucleus or cytoplasm, or both, they engage in every step of the posttranscriptional modification process, including alternative splicing, regulation of mRNA levels, transport between cellular compartments, alternative polyadenylation, transcript stability, etc. The TIAR protein has been shown to be transported from the nucleus to the cytoplasm during Fas-mediated apoptotic cell death [3]. Cytoplasmic RBPs, on the other hand, include Unr, which has been shown to be required for internally initiating the translation of human rhinovirus RNA [5]
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