Abstract
During mRNA transcription, diverse RNA-binding proteins (RBPs) are recruited to RNA polymerase II (RNAP II) transcription machinery. These RBPs bind to distinct sites of nascent RNA to co-transcriptionally operate mRNA processing. Recent studies have revealed a close relationship between transcription and co-transcriptional RNA processing, where one affects the other’s activity, indicating an essential role of protein–RNA interactions for the fine-tuning of mRNA production. Owing to their limited amount in cells, the detection of protein–RNA interactions specifically assembled on the transcribing RNAP II machinery still remains challenging. Currently, cross-linking and immunoprecipitation (CLIP) has become a standard method to detect in vivo protein–RNA interactions, although it requires a large amount of input materials. Several improved methods, such as infrared-CLIP (irCLIP), enhanced CLIP (eCLIP), and target RNA immunoprecipitation (tRIP), have shown remarkable enhancements in the detection efficiency. Furthermore, the utilization of an RNA editing mechanism or proximity labeling strategy has achieved the detection of faint protein–RNA interactions in cells without depending on crosslinking. This review aims to explore various methods being developed to detect endogenous protein–RNA interaction sites and discusses how they may be applied to the analysis of co-transcriptional RNA processing.
Highlights
Following transcription by RNA polymerase II (RNAP II), nascent RNA undergoes processing events of a 7-methyl guanosine cap at the 50 end (50 capping), splicing of exons, and formation of the 30 end by cleavage and polyadenylation, to produce mature mRNA (Figure 1)
This review aims to explore various methods being developed to detect endogenous protein–RNA interaction sites and discusses how they may be applied to the analysis of co-transcriptional RNA processing
This review explores the development of protein-centric methods to identify protein–RNA interaction sites in endogenous RNA and discusses their potential application to the analysis of cotranscriptional RNA processing
Summary
Following transcription by RNA polymerase II (RNAP II), nascent RNA undergoes processing events of a 7-methyl guanosine cap at the 50 end (50 capping), splicing of exons, and formation of the 30 end by cleavage and polyadenylation, to produce mature mRNA (Figure 1). The binding of splicing factors, such as U1 snRNP [3], SRSF2 [4], FUS [5], and SFPQ [6], to pre-mRNA affects transcription elongation in addition to the processing of nascent. The dynamic phosphorylation pattern of the C-terminal domain (CTD) of RNAP II plays an important role in the regulation of the transcriptional status of RNAP II [7], and active spliceosomes are complexed, especially to serine 5 phosphorylated CTD, during elongation [8]. This represents the physiological connection between RNAP II and mRNA processing. The transient nature of protein–RNA interactions complicates their experimental identification
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
