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bioRxiv · 10.1101/246827

NET-prism enables RNA polymerase-dedicated transcriptional interrogation at nucleotide resolution

Mylonas, C.·Tessarz, P.
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Abstract

The advent of quantitative approaches that enable interrogation of transcription at single nucleotide resolution has allowed a novel understanding of transcriptional regulation previously undefined. To better map transcription genome-wide at base pair resolution and with transcription/elongation factor dependency we developed an adapted NET-seq protocol called NET-prism (Native Elongating Transcription by Polymerase-Regulated Immunoprecipitants in the Mammalian genome). NET-prism introduces an immunoprecipitation to capture RNA Pol II - associated proteins, which reveals the interaction of these proteins with active RNA Pol II. Application of NET-prism on different Pol II subunits (Pol II S2ph, Pol II S5ph), elongation factors (Spt6, Ssrp1), and components of the pre-initiation complex (PIC) (TFIID, TBP, and Mediator) reveals diverse Pol II signals, at a single nucleotide resolution, with regards to directionality and intensity over promoters, splice sites, and enhancers/super-enhancers. NET-prism will be broadly applicable as it exposes transcription factor/Pol II dependent topographic specificity and thus, a new degree of regulatory complexity.

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Mylonas, C., Tessarz, P.. 2018-01-11. NET-prism enables RNA polymerase-dedicated transcriptional interrogation at nucleotide resolution. https://doi.org/10.1101/246827

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