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Key features of the interaction between Pcf11 CID and RNA polymerase II CTD

Nature Structural & Molecular Biology volume 12, pages 144–151 (2005)Cite this article

Abstract

The C-terminal domain (CTD) of the large subunit of RNA polymerase II is a platform for mRNA processing factors and links gene transcription to mRNA capping, splicing and polyadenylation. Pcf11, an essential component of the mRNA cleavage factor IA, contains a CTD-interaction domain that binds in a phospho-dependent manner to the heptad repeats within the RNA polymerase II CTD. We show here that the phosphorylated CTD exists as a dynamic disordered ensemble in solution and, by induced fit, it assumes a structured conformation when bound to Pcf11. In addition, we detected cis-trans populations for the CTD prolines, and found that only the all-trans form is selected for binding. These data suggest that the recognition of the CTD is regulated by independent site-specific modifications (phosphorylation and proline cis-trans isomerization) and, probably, by the local concentration of suitable binding sites.

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Figure 1: Peptides used in this study.
Figure 2: Conformation of CTD-derived peptides.
Figure 3: Cis-trans isomerization in CTD-derived peptides.
Figure 4: Structure of Pcf11 CID.
Figure 6: Binding of Pcf11 CID to CTD-derived peptides.
Figure 5: CID-PTSPSYSpPTSPSY interaction by ITC.
Figure 7: Cis-trans proline isomerization and CID binding.

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Acknowledgements

We thank S. Gamblin for assistance with crystal handling and critical reading of the manuscript. We also thank T. Frenkiel for assistance in recording NMR experiments, P. Fletcher for synthesizing the (YSpPTSPS)2 and (YSpPTSPS)3 peptides, and P. Temussi for sharing his expertise on studies of peptides.

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Author notes

  1. Christian G Noble and David Hollingworth: These authors contributed equally to this work.

Authors and Affiliations

  1. Division of Protein Structure, National Institute for Medical Research, The Ridgeway, Mill Hill, London, NW7 1AA, UK

    Christian G Noble, Valerie Ennis-Adeniran, Stephen J Smerdon & Ian A Taylor

  2. Division of Molecular Structure, National Institute for Medical Research, The Ridgeway, Mill Hill, London, NW7 1AA, UK

    David Hollingworth & Andres Ramos

  3. Division of Physical Biochemistry, National Institute for Medical Research, The Ridgeway, Mill Hill, London, NW7 1AA, UK

    Stephen R Martin

  4. Biomedical NMR Centre, National Institute for Medical Research, The Ridgeway, Mill Hill, London, NW7 1AA, UK

    Geoff Kelly

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Correspondence to Ian A Taylor or Andres Ramos.

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Supplementary information

Supplementary Fig. 1

Free peptide's ROESY. (PDF 91 kb)

Supplementary Fig. 2

Free peptide's 13C HSQC. (PDF 137 kb)

Supplementary Fig. 3

15N HSQC of Pcf11 constructs. (PDF 351 kb)

Supplementary Fig. 4

15N HSQC of Pcf11 titration. (PDF 276 kb)

Supplementary Fig. 5

Tr-NOE experiments. (PDF 203 kb)

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Noble, C., Hollingworth, D., Martin, S. et al. Key features of the interaction between Pcf11 CID and RNA polymerase II CTD. Nat Struct Mol Biol 12, 144–151 (2005). https://doi.org/10.1038/nsmb887

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