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Multisite phosphorylation networks as signal processors for Cdk1

Nature Structural & Molecular Biology volume 20, pages 1415–1424 (2013)Cite this article

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Abstract

The order and timing of cell-cycle events is controlled by changing substrate specificity and different activity thresholds of cyclin-dependent kinases (CDKs). However, it is not understood how a single protein kinase can trigger hundreds of switches in a sufficiently time-resolved fashion. We show that cyclin–Cdk1–Cks1–dependent phosphorylation of multisite targets in Saccharomyces cerevisiae is controlled by key substrate parameters including distances between phosphorylation sites, distribution of serines and threonines as phosphoacceptors and positioning of cyclin-docking motifs. The component mediating the key interactions in this process is Cks1, the phosphoadaptor subunit of the cyclin–Cdk1–Cks1 complex. We propose that variation of these parameters within networks of phosphorylation sites in different targets provides a wide range of possibilities for differential amplification of Cdk1 signals, thus providing a mechanism to generate a wide range of thresholds in the cell cycle.

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Figure 1: Cks1-dependent multisite phosphorylation of Cdk1 targets.
Figure 2: Exclusive preference of threonine over serine residues as the priming sites for Cks1-dependent docking and phosphorylation steps.
Figure 3: Analysis of the influence of distance between the priming phosphorylation site and the secondary phosphorylation site.
Figure 4: The optimal distances between the phosphorylation sites of Sic1 are critical for its degradation in vivo.
Figure 5: Analysis of the influence of distance between the phosphorylation site and the cyclin-specific docking sites.
Figure 6: Analysis of the processivity of multiphosphorylation.
Figure 7: Cks1 differentially stimulates the phosphorylation of various sites in Stb1, Ndd1 and Swi5.

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References

  1. Morgan, D.O. The Cell Cycle: Principles of Control (New Science Press, 2007).

  2. Ubersax, J.A. et al. Targets of the cyclin-dependent kinase Cdk1. Nature 425, 859–864 (2003).

    Article  CAS  Google Scholar 

  3. Holt, L.J. et al. Global analysis of Cdk1 substrate phosphorylation sites provides insights into evolution. Science 325, 1682–1686 (2009).

    Article  CAS  Google Scholar 

  4. Errico, A., Deshmukh, K., Tanaka, Y., Pozniakovsky, A. & Hunt, T. Identification of substrates for cyclin dependent kinases. Adv. Enzyme Regul. 50, 375–399 (2010).

    Article  Google Scholar 

  5. Pagliuca, F.W. et al. Quantitative proteomics reveals the basis for the biochemical specificity of the cell-cycle machinery. Mol. Cell 43, 406–417 (2011).

    Article  CAS  Google Scholar 

  6. Enserink, J.M. & Kolodner, R.D. An overview of Cdk1-controlled targets and processes. Cell Div. 5, 11 (2010).

    Article  Google Scholar 

  7. Stern, B. & Nurse, P. A quantitative model for the cdc2 control of S phase and mitosis in fission yeast. Trends Genet. 12, 345–350 (1996).

    Article  CAS  Google Scholar 

  8. Coudreuse, D. & Nurse, P. Driving the cell cycle with a minimal CDK control network. Nature 468, 1074–1079 (2010).

    Article  CAS  Google Scholar 

  9. Uhlmann, F., Bouchoux, C. & Lopez-Aviles, S. A quantitative model for cyclin-dependent kinase control of the cell cycle: revisited. Phil. Trans. R. Soc. Lond. B 366, 3572–3583 (2011).

    Article  CAS  Google Scholar 

  10. Fisher, D., Krasinska, L., Coudreuse, D. & Novak, B. Phosphorylation network dynamics in the control of cell cycle transitions. J. Cell Sci. 125, 4703–4711 (2012).

    Article  CAS  Google Scholar 

  11. Fisher, D.L. & Nurse, P. A single fission yeast mitotic cyclin B p34cdc2 kinase promotes both S-phase and mitosis in the absence of G1 cyclins. EMBO J. 15, 850–860 (1996).

    Article  CAS  Google Scholar 

  12. Songyang, Z. et al. Use of an oriented peptide library to determine the optimal substrates of protein kinases. Curr. Biol. 4, 973–982 (1994).

    Article  CAS  Google Scholar 

  13. Mok, J. et al. Deciphering protein kinase specificity through large-scale analysis of yeast phosphorylation site motifs. Sci. Signal. 3, ra12 (2010).

    Article  Google Scholar 

  14. Kõivomägi, M. et al. Dynamics of Cdk1 substrate specificity during the cell cycle. Mol. Cell 42, 610–623 (2011).

    Article  Google Scholar 

  15. Deibler, R.W. & Kirschner, M.W. Quantitative reconstitution of mitotic CDK1 activation in somatic cell extracts. Mol. Cell 37, 753–767 (2010).

    Article  CAS  Google Scholar 

  16. Gavet, O. & Pines, J. Progressive activation of cyclinB1-Cdk1 coordinates entry to mitosis. Dev. Cell 18, 533–543 (2010).

    Article  CAS  Google Scholar 

  17. Oikonomou, C. & Cross, F.R. Rising cyclin-CDK levels order cell cycle events. PLoS ONE 6, e20788 (2011).

    Article  CAS  Google Scholar 

  18. Cross, F.R., Yuste-Rojas, M., Gray, S. & Jacobson, M.D. Specialization and targeting of B-type cyclins. Mol. Cell 4, 11–19 (1999).

    Article  CAS  Google Scholar 

  19. Cross, F.R. & Jacobson, M.D. Conservation and function of a potential substrate-binding domain in the yeast Clb5 B-type cyclin. Mol. Cell Biol. 20, 4782–4790 (2000).

    Article  CAS  Google Scholar 

  20. Loog, M. & Morgan, D.O. Cyclin specificity in the phosphorylation of cyclin-dependent kinase substrates. Nature 434, 104–108 (2005).

    Article  CAS  Google Scholar 

  21. Bloom, J. & Cross, F.R. Multiple levels of cyclin specificity in cell-cycle control. Nat. Rev. Mol. Cell Biol. 8, 149–160 (2007).

    Article  CAS  Google Scholar 

  22. Kõivomägi, M. et al. Cascades of multisite phosphorylation control Sic1 destruction at the onset of S phase. Nature 480, 128–131 (2011).

    Article  Google Scholar 

  23. Kõivomägi, M. & Loog, M. Cdk1: a kinase with changing substrate specificity. Cell Cycle 10, 3625–3626 (2011).

    Article  Google Scholar 

  24. Mendenhall, M.D. & Hodge, A.E. Regulation of Cdc28 cyclin-dependent protein kinase activity during the cell cycle of the yeast Saccharomyces cerevisiae. Microbiol. Mol. Biol. Rev. 62, 1191–1243 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Kim, S.Y. & Ferrell, J.E. Jr. Substrate competition as a source of ultrasensitivity in the inactivation of Wee1. Cell 128, 1133–1145 (2007).

    Article  CAS  Google Scholar 

  26. Trunnell, N.B., Poon, A.C., Kim, S.Y. & Ferrell, J.E. Jr. Ultrasensitivity in the regulation of Cdc25C by Cdk1. Mol. Cell 41, 263–274 (2011).

    Article  CAS  Google Scholar 

  27. Strickfaden, S.C. et al. A mechanism for cell-cycle regulation of MAP kinase signaling in a yeast differentiation pathway. Cell 128, 519–531 (2007).

    Article  CAS  Google Scholar 

  28. Wagner, M.V. et al. Whi5 regulation by site specific CDK-phosphorylation in Saccharomyces cerevisiae. PLoS ONE 4, e4300 (2009).

    Article  Google Scholar 

  29. Burke, J.R., Hura, G.L. & Rubin, S.M. Structures of inactive retinoblastoma protein reveal multiple mechanisms for cell cycle control. Genes Dev. 26, 1156–1166 (2012).

    Article  CAS  Google Scholar 

  30. Harvey, S.L., Charlet, A., Haas, W., Gygi, S.P. & Kellogg, D.R. Cdk1-dependent regulation of the mitotic inhibitor Wee1. Cell 122, 407–420 (2005).

    Article  CAS  Google Scholar 

  31. Verma, R., Feldman, R.M. & Deshaies, R.J. SIC1 is ubiquitinated in vitro by a pathway that requires CDC4, CDC34, and cyclin/CDK activities. Mol. Biol. Cell 8, 1427–1437 (1997).

    Article  CAS  Google Scholar 

  32. Hao, B., Oehlmann, S., Sowa, M.E., Harper, J.W. & Pavletich, N.P. Structure of a Fbw7-Skp1-cyclin E complex: multisite-phosphorylated substrate recognition by SCF ubiquitin ligases. Mol. Cell 26, 131–143 (2007).

    Article  CAS  Google Scholar 

  33. Hayles, J., Beach, D., Durkacz, B. & Nurse, P. The fission yeast cell cycle control gene cdc2: isolation of a sequence suc1 that suppresses cdc2 mutant function. Mol. Gen. Genet. 202, 291–293 (1986).

    Article  CAS  Google Scholar 

  34. Hadwiger, J.A., Wittenberg, C., Richardson, H.E., de Barros Lopes, M. & Reed, S.I. A family of cyclin homologs that control the G1 phase in yeast. Proc. Natl. Acad. Sci. USA 86, 6255–6259 (1989).

    Article  CAS  Google Scholar 

  35. Tang, Y. & Reed, S.I. The Cdk-associated protein Cks1 functions both in G1 and G2 in Saccharomyces cerevisiae. Genes Dev. 7, 822–832 (1993).

    Article  CAS  Google Scholar 

  36. Kito, K., Kawaguchi, N., Okada, S. & Ito, T. Discrimination between stable and dynamic components of protein complexes by means of quantitative proteomics. Proteomics 8, 2366–2370 (2008).

    Article  CAS  Google Scholar 

  37. Patra, D., Wang, S.X., Kumagai, A. & Dunphy, W.G. The Xenopus Suc1/Cks protein promotes the phosphorylation of G2/M regulators. J. Biol. Chem. 274, 36839–36842 (1999).

    Article  CAS  Google Scholar 

  38. Patra, D. & Dunphy, W.G. Xe-p9, a Xenopus Suc1/Cks protein, is essential for the Cdc2-dependent phosphorylation of the anaphase-promoting complex at mitosis. Genes Dev. 12, 2549–2559 (1998).

    Article  CAS  Google Scholar 

  39. Reynard, G.J., Reynolds, W., Verma, R. & Deshaies, R.J. Cks1 is required for G1 cyclin-cyclin-dependent kinase activity in budding yeast. Mol. Cell Biol. 20, 5858–5864 (2000).

    Article  CAS  Google Scholar 

  40. Bourne, Y. et al. Crystal structure and mutational analysis of the Saccharomyces cerevisiae cell cycle regulatory protein Cks1: implications for domain swapping, anion binding and protein interactions. Structure 8, 841–850 (2000).

    Article  CAS  Google Scholar 

  41. Nash, P. et al. Multisite phosphorylation of a CDK inhibitor sets a threshold for the onset of DNA replication. Nature 414, 514–521 (2001).

    Article  CAS  Google Scholar 

  42. McGrath, D.A. et al. Cks confers specificity to phosphorylation-dependent Cdk signaling pathways. Nat. Struct. Mol. Biol. 10.1038/nsmb.2707 (3 November 2013).

  43. Orlicky, S., Tang, X., Willems, A., Tyers, M. & Sicheri, F. Structural basis for phosphodependent substrate selection and orientation by the SCFCdc4 ubiquitin ligase. Cell 112, 243–256 (2003).

    Article  CAS  Google Scholar 

  44. Brocca, S. et al. Order propensity of an intrinsically disordered protein, the cyclin-dependent-kinase inhibitor Sic1. Proteins 76, 731–746 (2009).

    Article  CAS  Google Scholar 

  45. Schulman, B.A., Lindstrom, D.L. & Harlow, E. Substrate recruitment to cyclin-dependent kinase 2 by a multipurpose docking site on cyclin A. Proc. Natl. Acad. Sci. USA 95, 10453–10458 (1998).

    Article  CAS  Google Scholar 

  46. Brown, N.R., Noble, M.E., Endicott, J.A. & Johnson, L.N. The structural basis for specificity of substrate and recruitment peptides for cyclin-dependent kinases. Nat. Cell Biol. 1, 438–443 (1999).

    Article  CAS  Google Scholar 

  47. Takeda, D.Y., Wohlschlegel, J.A. & Dutta, A. A bipartite substrate recognition motif for cyclin-dependent kinases. J. Biol. Chem. 276, 1993–1997 (2001).

    Article  CAS  Google Scholar 

  48. Wohlschlegel, J.A., Dwyer, B.T., Takeda, D.Y. & Dutta, A. Mutational analysis of the Cy motif from p21 reveals sequence degeneracy and specificity for different cyclin-dependent kinases. Mol. Cell Biol. 21, 4868–4874 (2001).

    Article  CAS  Google Scholar 

  49. Wilmes, G.M. et al. Interaction of the S-phase cyclin Clb5 with an “RXL” docking sequence in the initiator protein Orc6 provides an origin-localized replication control switch. Genes Dev. 18, 981–991 (2004).

    Article  CAS  Google Scholar 

  50. Lowe, E.D. et al. Specificity determinants of recruitment peptides bound to phospho-CDK2/cyclin A. Biochemistry 41, 15625–15634 (2002).

    Article  CAS  Google Scholar 

  51. Bhaduri, S. & Pryciak, P.M. Cyclin-specific docking motifs promote phosphorylation of yeast signaling proteins by G1/S Cdk complexes. Curr. Biol. 21, 1615–1623 (2011).

    Article  CAS  Google Scholar 

  52. Bourne, Y. et al. Crystal structure and mutational analysis of the human CDK2 kinase complex with cell cycle-regulatory protein CksHs1. Cell 84, 863–874 (1996).

    Article  CAS  Google Scholar 

  53. Bouchoux, C. & Uhlmann, F. A quantitative model for ordered Cdk substrate dephosphorylation during mitotic exit. Cell 147, 803–814 (2011).

    Article  CAS  Google Scholar 

  54. Hirschi, A. et al. An overlapping kinase and phosphatase docking site regulates activity of the retinoblastoma protein. Nat. Struct. Mol. Biol. 17, 1051–1057 (2010).

    Article  CAS  Google Scholar 

  55. Ikui, A.E., Rossio, V., Schroeder, L. & Yoshida, S. A yeast GSK-3 kinase Mck1 promotes Cdc6 degradation to inhibit DNA re-replication. PLoS Genet. 8, e1003099 (2012).

    Article  CAS  Google Scholar 

  56. Lyons, N.A., Fonslow, B.R., Diedrich, J.K., Yates, J.R. III & Morgan, D.O. Sequential primed kinases create a damage-responsive phosphodegron on Eco1. Nat. Struct. Mol. Biol. 20, 194–201 (2013).

    Article  CAS  Google Scholar 

  57. Puig, O. et al. The tandem affinity purification (TAP) method: a general procedure of protein complex purification. Methods 24, 218–229 (2001).

    Article  CAS  Google Scholar 

  58. McCusker, D. et al. Cdk1 coordinates cell-surface growth with the cell cycle. Nat. Cell Biol. 9, 506–515 (2007).

    Article  CAS  Google Scholar 

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Acknowledgements

We would like to thank L. Holt and D. Morgan for valuable comments on the manuscript. We thank D. Kellogg (University of California, Santa Cruz) for kindly providing the strains and J. Mihhejev for excellent technical assistance and S. Kasvandik for assistance with MS. This work was supported by an International Senior Research Fellowship from the Wellcome Trust (no. 079014/Z/06/Z , to M.L.), an installation grant from the European Molecular Biology Organization and Howard Hughes Medical Institute (no. 1253, to M.L.), a targeted financing scheme and an institutional grant from the Estonian government (IUT2-21, to M.L.) and funding to S.M.R. from the American Cancer Society (RSG-12-131-01-CCG).

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Authors and Affiliations

  1. Institute of Technology, University of Tartu, Tartu, Estonia

    Mardo Kõivomägi, Mihkel Örd, Anna Iofik, Ervin Valk, Rainis Venta, Ilona Faustova, Rait Kivi & Mart Loog

  2. Department of Molecular, Cell, and Developmental Biology, University of California, Santa Cruz, Santa Cruz, California, USA

    Eva Rose M Balog

  3. Department of Chemistry and Biochemistry, University of California, Santa Cruz, Santa Cruz, California, USA

    Seth M Rubin

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Contributions

M.K., M.Ö., A.I., E.V., R.V., I.F. and R.K. designed and performed the experiments. The ITC experiments were performed by E.R.M.B., and the structural model was constructed by S.M.R.; M.L. coordinated the project and wrote the manuscript with assistance from S.M.R., M.K. and M.Ö.

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Correspondence to Mart Loog.

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Integrated supplementary information

Supplementary Figure 1 Analysis of the Cdk1-dependent phosphorylation of Sic1ΔC version with the threonines at the CDK phosphorylation sites mutated to serines, and additional control experiments supplementing the analysis of Cks1-dependent phosphorylation steps using the two-site Sic1ΔC-based substrate constructs.

(a,b) The comparison of the phosphorylation time courses of the wild type Sic1ΔC and all-Ser-Sic1ΔC using Cln2-Cdk1-Cks1 (a) and Clb5-Cdk1-Cks1 (b); (c) The turnover of primary substrates (S0) for the experiment presented in Fig. 2b; (d) Multisite phosphorylation patterns of wild type Sic1ΔC and all-Ser-Sic1ΔC in reaction with Cks1wt and Cks1mut using Clb2-Cdk1-Cks1 complex; (e) Phosphorylation of two-site Sic1ΔC-based substrate constructs containing the indicated sites confirms that phosphorylation of secondary site T48 requires a threonine-based N-terminal priming site; (f) A control experiment to supplement Fig. 3b shows that the suboptimal secondary sites at positions +12 and +14 are not likely to be phosphorylated independently of the priming site; (g) A control experiment to supplement Fig. 6b-e confirms that the two-site construct T5-T33-Sic1ΔC can be used for the processivity experiments as the serine version (S5-S33) of the construct did not accumulate the two-phosphate form comparably to T5-T33-Sic1ΔC in the assay conditions and the reaction time-window used.

Supplementary Figure 2 The in vivo demonstration of the requirement of threonine as the N-terminal priming site for Cks1-dependent phosphorylation steps.

(a,b) The intermediate phosphorylated forms of the all-serine version of Sic1 accumulate in vivo. The overexpression of the all-Ser-Sic1 caused inviability of cells as shown in Fig. 2d. In order to demonstrate that the protein levels and the phosphorylation shifts are different in case of wild type Sic1 and all-ser-Sic1 mutant, the phosphorylation patterns were followed after the induction of the Sic1 versions by galactose in hydroxyurea (a) or nocodazole (b) arrested cells; (c) Cells constitutively expressing the Sic1ΔC-based constructs containing either threonine or serine in position 5 were released from a pheromone arrest and the phosphorylation shifts were analyzed using Phos-Tag SDS-PAGE and western blotting. The two lower panels present similar experiments with the exception that the cyclin-specific docking motifs were mutated in the Sic1ΔC constructs (rxl-vllpp mutation14).

Supplementary Figure 3 The proline in position –2 is an additional specificity determinant for phosphoepitope interaction with Cks1.

(a) The effect of -2Pro on the rate of secondary phosphorylation was studied using the substrate constructs T33-T48-Sic1ΔC and -2P-T33-T48-Sic1ΔC; (b) Full-length version of Sic1 containing the Pro in position -2 from the site T33 was overexpressed under the galactose promoter to assay the ability of cells to degrade the construct T33-T45-T48-S76-S80-Sic1; (c) Representative ITC data for Cks1/phospho-Sic1 binding experiments. The values of the obtained binding constants for the indicated Sic1ΔC-based constructs are given in Supplementary Table 1.

Supplementary Figure 4 Relative effects of cyclin-docking sites and Cdk consensus sites on the Cks1-dependent phosphorylation steps and analysis of the distance requirements for a Cks1-dependent docking step.

(a-c) The effect of the hpm mutation and the mutation of lysine in position +3 on the rate of the secondary phosphorylation at position T33 was studied using the constructs T5-T33-Sic1ΔC and T5-T33+3A-Sic1ΔC; (d) Diagram showing the distance requirement for Cks1-dependent docking steps studied using Sic1ΔC-based substrate constructs containing T5 as the primary phosphorylation site and suboptimal TP sites as secondary sites using Clb5- and Cln2-Cdk1; (e-g) A scheme and examples of MS spectra of the analysis of distance requirements for Cks1-dependnet phosphorylation step in case of Far1; (h,i) The effect of the replacement of optimal CDK consensus motif with a suboptimal motif on the phosphorylation of the indicated Sic1ΔC-based substrate constructs; (j) The suboptimal Cdk1 sites (SP and TP consensus sites) do not get phosphorylated in vivo without the help of Cks1- or cyclin-dependent docking interactions.

Supplementary Figure 5 A general kinetic scheme of a sequential multiphosphorylation process.

For the sake of clarity, each form of substrate containing a certain number of phosphates is considered as kinetically equal.

Supplementary Figure 6 Modeling of processivity in the phosphorylation of a two-site kinase substrate.

(a) The kinetic scheme outlining the phosphorylation events. (b) The ODE model used for simulation of the time courses. (c-h) The time course simulations of the ODE model.

Supplementary Figure 7 Processivity analysis of the phosphorylation of Sic1ΔC-based constructs Whi5 and Fin1.

(a) Schemes showing the positions of phosphorylation sites in substrate constructs used in the experiments presented in panels 'b-d' (b) Experiment with low enzyme concentrations, similar to the ones presented in Fig. 6d,e, performed for T33+12TP-Sic1ΔC; (c) Full time courses of phosphorylation of T5-T33-Sic1ΔC with Clb5-Cdk1 in complex with Cks1wt and Cks1mut; (d) Graph showing the XMP values obtained by the quantification of data from panel 'c'. (e-j) The analysis of the processivity of Whi5 and Fin1 phosphorylation. A series of low concentrations of cyclin-Cdk1 complexes were used in a similar assay as presented for the Sic1ΔC-based constructs in Figure 6.

Supplementary Figure 8 Uncropped lanes from the autoradiography scans and western blots.

The horizontal panels 'a' and 'b' belong to Fig.1, panels 'd-f' to Fig.2, panels 'g-i' to Fig. 3, panels 'j' and 'k' to Fig. 6. The coomassie stained Phos-Tag gels in panel 'c' show the estimated migration of unphosphorylated or phosphorylated Sic1ΔC containing single Cdk1 site (Sic1ΔC-T33, Sic1ΔC-T33S) or all physiological phosphorylation sites (Sic1ΔC-wt). The arrows indicate the approximate estimated migration of nonradioactive unphosphorylated protein. For the rest of the details see the Online Methods and the Supplementary Results section.

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Kõivomägi, M., Örd, M., Iofik, A. et al. Multisite phosphorylation networks as signal processors for Cdk1. Nat Struct Mol Biol 20, 1415–1424 (2013). https://doi.org/10.1038/nsmb.2706

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