A multisubunit factor, CstF, is required for polyadenylation of mammalian pre-mRNAs

Y. Takagaki, J. L. Manley, C. C. MacDonald, J. Wilusz, T. Shenk

Research output: Contribution to journalArticle

175 Scopus citations

Abstract

We have purified and characterized a factor required for accurate polyadenylation of mammalian pre-mRNAs in vitro. This factor, called cleavage-stimulation factor (CstF), is composed of three distinct polypeptide subunits of 77, 64, and 50 kD. Using monoclonal antibodies directed against the 64- and 50-kD subunits, we show that CstF is required for efficient cleavage of polyadenylation substrates. Furthermore, CstF present in unfractionated nuclear extracts interacts with pre-mRNAs containing the signal sequence AAUAAA, but not AAGAAA, in such a manner that the 64-kD subunit can be cross-linked to the RNA by UV light. This polypeptide is thus identical to the previously described 64-kD nuclear protein that binds to AAUAAA-polypeptide is thus identical to the previously described 64-kD nuclear protein that binds to AAUAAA-containing RNAs. Finally, indirect immunofluorescence of fixed cells indicates that CstF is distributed diffusely throughout the nucleus in a granular pattern distinct from the 'speckled' pattern displayed by factors involved in pre-mRNA splicing, but similar to that of heterogeneous nuclear ribonucleoproteins. A model is presented in which CstF binds specifically to nascent RNA polymerase II transcripts and, by interacting with other factors, results in a rapid initiation of 3'-end processing of pre-mRNAs.

Original languageEnglish (US)
Pages (from-to)2112-2120
Number of pages9
JournalGenes and Development
Volume4
Issue number12 A
DOIs
StatePublished - 1990

All Science Journal Classification (ASJC) codes

  • Genetics
  • Developmental Biology

Keywords

  • Multisubunit factor CstF
  • mRNA processing
  • polyadenylation

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