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Splicing- and cleavage-independent requirement of RNA polymerase II CTD for mRNA release from the transcription site
Correspondence to Maria Carmo-Fonseca: carmo.fonseca{at}fm.ul.pt
Eukaryotic cells have a surveillance mechanism that identifies aberrantly processed pre-mRNAs and prevents their flow to the cytoplasm by tethering them near the site of transcription. Here we provide evidence that mRNA release from the transcription site requires the heptad repeat structure of the C-terminal domain (CTD) of RNA polymerase II. The mammalian CTD, which is essential for normal co-transcriptional maturation of mRNA precursors, comprises 52 heptad repeats. We show that a truncated CTD containing 31 repeats (heptads 1–23, 36–38, and 48–52) is sufficient to support transcription, splicing, cleavage, and polyadenylation. Yet, the resulting mRNAs are mostly retained in the vicinity of the gene after transcriptional shutoff. The retained mRNAs maintain the ability to recruit components of the exon junction complex and the nuclear exosome subunit Rrp6p, suggesting that binding of these proteins is not sufficient for RNA release. We propose that the missing heptads in the truncated CTD mutant are required for binding of proteins implicated in a final co-transcriptional maturation of spliced and 3' end cleaved and polyadenylated mRNAs into export-competent ribonucleoprotein particles.
Abbreviations used in this paper: ßLCR, HBB micro-locus control region; CMV, cytomegalovirus; CTD, C-terminal domain; EJC, exon junction complex; HA, haemagglutinin; HBB, human ß-globin gene; Hbb1, murine ßmajor-globin gene; MEL, murine erythroleukemia; RNA Pol II LS, largest subunit of RNA polymerase II.
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