Elongation factor-specific capture of RNA polymerase II complexes
| Autor: | Lea H. Gregersen, Richard Mitter, Jesper Q. Svejstrup |
|---|---|
| Rok vydání: | 2023 |
| Předmět: |
mNET-seq
termination Model organisms Chemical Biology & High Throughput nascent transcription SCAF8 Genome Integrity & Repair Gene Expression SCAF4 Tumour Biology Biochemistry & Proteomics Biochemistry Genetics and Molecular Biology (miscellaneous) Biochemistry Computer Science Applications molecular biology [CP] Genetics Radiology Nuclear Medicine and imaging RNA polymerase II RNAPII RNAPII transcript elongation transcription Genetics & Genomics RNAPII CTD-associated factors Biotechnology Computational & Systems Biology |
| Zdroj: | Gregersen, L H, Mitter, R & Svejstrup, J Q 2022, ' Elongation factor-specific capture of RNA polymerase II complexes ', Cell Reports Methods, vol. 2, no. 12, 100368 . https://doi.org/10.1016/j.crmeth.2022.100368 |
| DOI: | 10.25418/crick.21814755.v1 |
| Popis: | Transcription of protein-coding genes is regulated by dynamic association of co-factors with RNA polymerase II (RNAPII). The function of these factors and their relationship with RNAPII is often poorly understood. Here, we present an approach for elongation-factor-specific mNET capture (ELCAP) of RNAPII complexes for sequencing and mass spectrometry analysis aimed at investigating the function of such RNAPII regulatory proteins. As proof of principle, we apply ELCAP to the RNAPII-associated proteins SCAF4 and SCAF8, which share an essential role as mRNA anti-terminators but have individual roles at the 3′ end of genes. Mass spectrometry analysis shows that both SCAF4 and SCAF8 are part of RNAPII elongation complexes containing 3′ end processing factors but depleted of splicing components. Importantly, the ELCAP sequencing (ELCAP-seq) profiles of SCAF4- and SCAF8-RNAPII complexes nicely reflect their function as mRNA-anti-terminators and their competing functions at the end of genes, where they prevent or promote transcriptional readthrough. |
| Databáze: | OpenAIRE |
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