POINT technology illuminates the processing of polymerase-associated intact nascent transcripts

Rui Sousa-Luís, Gwendal Dujardin, Inna Zukher, Hiroshi Kimura, Carika Weldon, Maria Carmo-Fonseca, Nick J. Proudfoot, Takayuki Nojima

Research output: Contribution to journalArticlepeer-review

4 Citations (Scopus)

Abstract

Mammalian chromatin is the site of both RNA polymerase II (Pol II) transcription and coupled RNA processing. However, molecular details of such co-transcriptional mechanisms remain obscure, partly because of technical limitations in purifying authentic nascent transcripts. We present a new approach to characterize nascent RNA, called polymerase intact nascent transcript (POINT) technology. This three-pronged methodology maps nascent RNA 5′ ends (POINT-5), establishes the kinetics of co-transcriptional splicing patterns (POINT-nano), and profiles whole transcription units (POINT-seq). In particular, we show by depletion of the nuclear exonuclease Xrn2 that this activity acts selectively on cleaved 5′ P-RNA at polyadenylation sites. Furthermore, POINT-nano reveals that co-transcriptional splicing either occurs immediately after splice site transcription or is delayed until Pol II transcribes downstream sequences. Finally, we connect RNA cleavage and splicing with either premature or full-length transcript termination. We anticipate that POINT technology will afford full dissection of the complexity of co-transcriptional RNA processing.

Original languageEnglish
Pages (from-to)1935-1950.e6
JournalMolecular Cell
Volume81
Issue number9
DOIs
Publication statusPublished - May 6 2021

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

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