Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Pol5p, a novel binding partner to Cdc10p in fission yeast involved in rRNA production

Abstract

Cdc10p is a major component of the cell cycle transcription factor complex MBF that controls G1–S phase specific gene expression in the fission yeast Schizosaccharomyces pombe. Here, we describe the identification of a new binding partner to Cdc10p and Pol5p. Pol5p was discovered through a 2-hybrid screen, with the direct interaction confirmed by in vitro “pull-down” experiments with bacterially expressed proteins. Pol5p appears to have no role in cell cycle gene expression, but is instead required for rRNA production. Pol5p is an essential gene, expressed constitutively throughout both the mitotic and meiotic life cycles, and localises to the nucleus. Over-expressing Pol5p has no phenotype, but reducing levels of Pol5p inhibits rRNA production. Pol5p is shown to bind to rDNA promoter fragments. Potentially, we have identified a mechanism by which Cdc10p controls rDNA gene expression, therefore linking the cell cycle with cellular growth.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

References

  1. Aves SJ, Durkacz BW, Carr A, Nurse P (1985) Cloning, sequencing and transcriptional control of the Schizosaccharomyces pombe cdc10 ‘start’ gene. EMBO J 4:457–463

  2. Bähler J (2005) Cell-cycle control of gene expression in budding and fission yeast. Annu Rev Genet 39:69–94

  3. Bähler J, Wu JQ, Longtine MS, Shah NG, McKenzie A, Steever AB, Wach A, Philippsen P, Pringle JR (1998) Heterologous modules for efficient and versatile PCR-based gene targeting in Schizosaccharomyces pombe. Yeast 14:943–951

  4. Breeden L (1996) Start-specific transcription in yeast. Curr Top Microbiol Immunol 208:95–127

  5. Breeden LL (2000) Cyclin transcription: timing is everything. Curr Biol 10:586–588

  6. Celenza JL, Carlson M (1989) Mutational analysis of the Saccharomyces cerevisiae SNF1 protein kinase and functional evidence for functional interaction with the SNF4 protein. Mol Cell Biol 9:5034–5044

  7. Cho RJ, Campbell MJ, Winzeler EA, Steinmetz L, Conway A, Wodicka L, Wolfsberg TG, Gabrielian AE, Landsman D, Lockhart DJ, Davis RW (1998) A genome-wide transcriptional analysis of the mitotic cell cycle. Mol Cell 2:65–73

  8. Cunliffe L, White S, McInerny CJ (2004) DSC1-MCB regulation of meiotic transcription in fission yeast. Mol Genet Genomics 271:60–70

  9. Feinberg AP, Volgelstein B (1984) A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem 1:6–13

  10. Gutz H, Heslot H, Leupold U, Loprieno N (1974) Schizosaccharomyces pombe. In: King RC (ed) Handbook of genetics, vol 1. Plenum, New York, pp 395–446

  11. Kohli J, Hottinger H, Munz P, Strauss A, Thuriaux P (1977) Genetic mapping in Schizosaccharomyces pombe by mitotic and meiotic analysis and induced haploidization. Genetics 87:23–471

  12. Lowndes NF, McInerny CJ, Johnson AL, Fantes PA, Johnston LH (1992) Control of DNA synthesis genes in fission yeast by the cell-cycle gene cdc10+. Nature 355:449–453

  13. Maundrell K (1993) Thiamine-repressible expression vectors pREP and pRIP for fission yeast. Gene 123:127–30

  14. McInerny CJ (2004) Cell cycle-regulated transcription in fission yeast. Biochem Soc Trans 32:967–972

  15. McInerny CJ, Kersey PJ, Creanor J, Fantes PA (1995) Positive and negative roles for cdc10 in cell cycle gene expression. Nucleic Acids Res 23:4761–4768

  16. Moreno S, Klar A, Nurse P (1991) Molecular genetic analysis of the fission yeast Schizosaccharomyces pombe. Meth Enzymol 194:795–823

  17. Ng SS, Anderson M, White S, McInerny CJ (2001) mik1+ G1-S transcription regulates mitotic entry in fission yeast. FEBS Lett 503:131–134

  18. Nurse P, Bissett Y (1981) Gene required in G1 for commitment to cell cycle and in G2 for control of mitosis in fission yeast. Nature 292:558–560

  19. Nurse P, Fantes P (1981) Cell cycle controls in fission yeast: a genetic analysis. In: John PCL (ed) The cell cycle. Cambridge University Press, Cambridge, pp 85–98

  20. Reymond A, Schmidt S, Simanis V (1992) Mutations in the cdc10 start gene of Schizosaccharomyces pombe implicate the region of homology between cdc10 and SWI6 as important for p85cdc10 function. Mol Gen Genet 234:449–456

  21. Reynolds N, Warbrick E, Fantes PA, MacNeill SA (2000) Essential interaction between the fission yeast DNA polymerase d subunit Cdc27 and Pcn1 (PCNA) mediated through a C-terminal p21Cip1-like PCNA binding motif. EMBO J 19:1108–1118

  22. Rustici G, Mata J, Kivinenm K, Lio P, Penkett CJ, Burns G, Hayles J, Brazma A, Nurse P, Bähler J (2004) Periodic gene expression program of the fission yeast cell cycle. Nat Genet 8:809–817

  23. Sambrook J, Fritch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

  24. Shorter J, Watson R, Giannakou ME, Clarke M, Warren G, Barr F (1999) GRASP55, a second mammalian GRASP protein involved in the stacking of golgi cisternae in a cell-free system. EMBO J 18:4949–4960

  25. Shimizu K, Kawasaki Y, Hiraga S, Tawaramoto M, Nakashima N, Sugino A (2002) The fifth essential DNA polymerase φ in Saccharomyces cerevisiae is localized to the nucleolus and plays an important role in synthesis of rRNA. Proc Nat Acad Sci USA 99:9133–9138

  26. Spellman PT, Sherlock G, Zhang MQ, Iyer VR, Anders K, Eisen MB, Brown PO, Botstein D, Futcher B (1998) Comprehensive identification of cell cycle-regulated genes of the yeast Saccharomyces cerevisiae by microarray hybridization. Mol Biol Cell 9:3273–3297

  27. Tavner FJ, Simpson R, Tashiro S, Favier D, Jenkins NA, Gilbert DJ, Copeland NG, Macmillan EM, Lutwyche J, Keough RA, Ishii S, Gonda TJ (1998) Molecular cloning reveals that the p160 Myb-binding protein is a novel, predominantly nucleolar protein which may play a role in transactivation by Myb. Mol Cell Biol 18:989–1002

  28. Wood V, Gwilliam R, Rajandream MA, Lyne M, Lyne R, Stewart A, Sgouros J, Peat N, Hayles J, Baker S, et al (2002) The genome sequence of Schizosaccharomyces pombe. Nature 415:871–880

  29. Yang W, Rogozin IB, Koonin EV (2003) Yeast POL5 is an evolutionarily conserved regulator of rDNA transcription unrelated to any known DNA polymerases. Cell Cycle 2:120–122

Download references

Acknowledgements

The work was supported by grants from the Wellcome Trust and the MRC. F.K.M and D.B were supported by studentships from the BBSRC and the Wellcome Trust. We thank Laura Gillies, Mridu Acharya and Andrew Hughes for assistance with the GFP and gel retardation experiments, and other members of the lab for help and encouragement during the course of this work.

Author information

Correspondence to Christopher J. McInerny.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Nadeem, F.K., Blair, D. & McInerny, C.J. Pol5p, a novel binding partner to Cdc10p in fission yeast involved in rRNA production. Mol Genet Genomics 276, 391–401 (2006). https://doi.org/10.1007/s00438-006-0144-6

Download citation

Keywords

  • Cell cycle
  • Fission yeast
  • MBF
  • Pol5p
  • rRNA