Coupled in vitro synthesis and splicing of RNA polymerase II transcripts

Sagarmoy Ghosh, Mariano A. Garcia-Blanco

Research output: Contribution to journalArticlepeer-review

54 Scopus citations

Abstract

Compelling in vivo studies suggest a tight functional linkage between RNA polymerase II transcription and pre-messenger RNA splicing. At present, the specific interactions involved in this coupling are poorly understood and deserve investigation. To this end, we developed an in vitro system that permits study of coupled transcription and splicing. Transcripts generated by RNA polymerase II were accurately and efficiently spliced under reaction conditions that permitted both transcription and splicing to occur simultaneously. The splicing of RNA-polymerase-II-driven transcripts was accelerated relative to that of the same transcripts driven by T7 RNA polymerase. Moreover, the product of exon ligation was found associated with the DNA template in reactions driven by RNA polymerase II. These two findings indicate that transcription and splicing were coupled in the in vitro system driven by RNA polymerase II, and suggest that this system will be useful for the biochemical study of this coupling.

Original languageEnglish (US)
Pages (from-to)1325-1334
Number of pages10
JournalRNA
Volume6
Issue number9
DOIs
StatePublished - 2000

Keywords

  • Cotranscriptional splicing
  • RNA polymerase II
  • Rate of splicing
  • T7 RNA polymerase
  • Transcription

ASJC Scopus subject areas

  • Molecular Biology

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