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CUL-2 is required for the G1-to-S-phase transition and mitotic chromosome condensation in Caenorhabditis elegans

Abstract

The human cullin protein CUL-2 functions in a ubiquitin-ligase complex with the von Hippel–Lindau (VHL) tumour suppressor protein. Here we show that, in Caenorhabditis elegans, cul-2 is expressed in proliferating cells and is required at two distinct points in the cell cycle, the G1-to-S-phase transition and mitosis. cul-2 mutant germ cells undergo a G1-phase arrest that correlates with accumulation of CKI-1, a member of the CIP/KIP family of cyclin-dependent-kinase inhibitors. In cul-2 mutant embryos, mitotic chromosomes are unable to condense, leading to unequal DNA segregation, chromosome bridging and the formation of multiple nuclei.

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Figure 1: Developmental expression of cul-2.
Figure 2: cul-2 deletion mutant.
Figure 3: G1 arrest of cul-2 germ cells.
Figure 4: cki-1 and cki-2 cloning and RNAi phenotype.
Figure 5: Level of CKI-1 protein in cul-2 mutants.
Figure 6: cul-2 mutant embryonic phenotype.
Figure 7: Cell-cycle lineage of wild-type and cul-2 mutant embryos.
Figure 8: Chromosome condensation is impaired in cul-2 mutant embryos.

References

  1. 1

    Pines, J. Cyclins and cyclin-dependent kinases: a biochemical view. Biochem. J. 308, 697–711 (1995).

    CAS  Article  Google Scholar 

  2. 2

    Hershko, A. & Ciechanover, A. The ubiquitin system. Annu. Rev. Biochem. 67, 425–479 (1998).

    CAS  Article  Google Scholar 

  3. 3

    Peters, J.-M. SCF and APC: the Yin and Yang of cell cycle regulated proteolysis. Curr. Opin. Cell Biol. 10, 759–768 (1998).

    CAS  Article  Google Scholar 

  4. 4

    Michael, W. M. & Newport, J. Coupling of mitosis to the completion of S phase through Cdc34-mediated degradation of Wee1. Science 282, 1886–1889 (1998).

    CAS  Article  Google Scholar 

  5. 5

    Kaiser, P., Sia, R. A. L., Bardes, E. G. S., Lew, D. J. & Reed, S. I. Cdc34 and the F-box protein Met30 are required for degradation of the cdk-inhibitory kinase Swe1. Genes Dev. 12, 2587–2597 (1998).

    CAS  Article  Google Scholar 

  6. 6

    Schwob, E., Bohm, T., Mendenhall, M. D. & Nasmyth, K. The B-type cyclin kinase inhibitor p40SIC1 controls the G1 to S transition in S. cerevisiae. Cell 79, 233–244 (1994).

    CAS  Article  Google Scholar 

  7. 7

    Feldman, R. M. R., Correll, C. C., Kaplan, K. B. & Deshaies, R. J. A complex of Cdc4p, Skp1p, and Cdc53p/Cullin catalyzes ubiquitination of the phosphorylated CDK inhibitor Sic1p. Cell 91, 221–230 (1997).

    CAS  Article  Google Scholar 

  8. 8

    Skowyra, D., Craig, K. L., Tyers, M., Elledge, S. J. & Harper, J. W. F-Box proteins are receptors that recruit phosphorylated substrates to the SCF ubiquitin-ligase complex. Cell 91, 209–219 (1997).

    CAS  Article  Google Scholar 

  9. 9

    Kamura, T. et al. Rbx1, a component of the VHL tumor suppressor complex and SCF ubiquitin ligase. Science 284, 657– 661 (1999).

    CAS  Article  Google Scholar 

  10. 10

    Skowyra, D. et al. Reconstitution of G1 cyclin ubiquitination with complexes containing SCFGrr1 and Rbx1. Science 284, 662–665 (1999).

    CAS  Article  Google Scholar 

  11. 11

    Ohta, T., Michel, J. J., Schottelius, A. J. & Xiong, Y. ROC1, a homolog of APC11, represents a family of cullin partners with an associated ubiquitin ligase activity. Mol. Cell 3, 535–541 (1999).

    CAS  Article  Google Scholar 

  12. 12

    Tan, P. et al. Recruitment of a ROC1-CUL1 ubiquitin ligase by Skp1 and HOS to catalyze the ubiquitination of IkBα. Mol. Cell 3, 527–533 (1999).

    CAS  Article  Google Scholar 

  13. 13

    Bai, C. et al. SKP1 connects cell cycle regulators to the ubiquitin proteolysis machinery through a novel motif, the F-box. Cell 86, 263–274 (1996).

    CAS  Article  Google Scholar 

  14. 14

    Deshaies, R. J., Chau, V. & Kirschner, M. Ubiquitination of the G1 cyclin Cln2p by a Cdc34p-dependent pathway. EMBO J. 14, 303– 312 (1995).

    CAS  Article  Google Scholar 

  15. 15

    Willems, A. R. et al. Cdc53 targets phosphorylated G1 cyclins for degradation by the ubiquitin proteolytic pathway. Cell 86, 453–463 (1996).

    CAS  Article  Google Scholar 

  16. 16

    Henchoz, S. et al. Phosphorylation- and ubiquitin-dependent degradation of the cyclin-dependent kinase inhibitor Far1p in budding yeast. Genes Dev. 11, 3046–3060 (1997).

    CAS  Article  Google Scholar 

  17. 17

    Kipreos, E. T., Lander, L. E., Wing, J. P., He, W. W. & Hedgecock, E. M. cul-1 is required for cell cycle exit in C. elegans and identifies a novel gene family. Cell 85, 829–839 (1996).

    CAS  Article  Google Scholar 

  18. 18

    Pause, A. et al. The von Hippel-Lindau tumor-suppressor gene product forms a stable complex with human CUL-2, a member of the Cdc53 family of proteins. Proc. Natl Acad. Sci. USA 94, 2155– 2161 (1997).

    Article  Google Scholar 

  19. 19

    Lonergan, K. M. et al. Regulation of hypoxia-inducible mRNAs by the von Hippel-Lindau tumor suppressor protein requires binding to complexes containing Elongins B/C and Cul2. Mol. Cell Biol. 18, 732– 741 (1998).

    CAS  Article  Google Scholar 

  20. 20

    Lisztwan, J., Imbert, G., Wirbelauer, C., Gstaiger, M. & Krek, W. The von Hippel-Lindau tumor suppressor protein is a component of an E3 ubiquitin-protein ligase activity. Genes Dev. 13, 1822–1833 (1999).

    CAS  Article  Google Scholar 

  21. 21

    Maxwell, P. H. et al. The tumour suppressor protein VHL targets hypoxia-inducible factors for oxygen-dependent proteolysis. Nature 399, 271–275 (1999).

    CAS  Article  Google Scholar 

  22. 22

    Kaelin, W. G. & Maher, E. R. The VHL tumour-suppressor gene paradigm. Trends Genet. 14, 423– 426 (1998).

    CAS  Article  Google Scholar 

  23. 23

    Hedgecock, E. M. & White, J. G. Polyploid tissues in the nematode Caenorhabditis elegans. Dev. Biol. 107, 128–133 (1985).

    CAS  Article  Google Scholar 

  24. 24

    Fire, A. et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391, 806–811 (1998).

    CAS  Article  Google Scholar 

  25. 25

    Hong, G., Roy, R. & Ambros, V. Developmental regulation of a cyclin-dependent kinase inhibitor controls postembryonic cell cycle progression in C. elegans. Development 125, 3585–3597 (1998).

    CAS  PubMed  Google Scholar 

  26. 26

    Mains, P. E., Kemphues, K. J., Sprunger, S. A., Sulston, I. A. & Wood, W. B. Mutations affecting the meiotic and mitotic divisions of the early Caenorhabditis elegans embryo. Genetics 126, 593–605 (1990).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. 27

    Edgar, L. G. & McGhee, J. D. DNA synthesis and the control of embryonic gene expression in C. elegans. Cell 53, 589–599 (1988).

    CAS  Article  Google Scholar 

  28. 28

    Hendzel, M. J. et al. Mitosis-specific phosphorylation of histone H3 initiates primarily within pericentromeric heterochromatin during G2 and spreads in an ordered fashion coincident with mitotic chromosome condensation. Chromosoma 106, 348–360 (1997).

    CAS  Article  Google Scholar 

  29. 29

    Wei, Y., Yu, L., Bowen, J., Gorovsky, M. A. & Allis, C. D. Phosphorylation of histone H3 is required for proper chromosome condensation and segregation. Cell 97, 99–109 (1999).

    CAS  Article  Google Scholar 

  30. 30

    Koshland, D. & Strunnikov, A. Mitotic chromosome condensation. Annu. Rev. Cell Dev. Biol. 12, 305– 333 (1996).

    CAS  Article  Google Scholar 

  31. 31

    Boxem, M., Srinivasan, D. G. & van den Heuvel, S. The Caenorhabditis elegans gene ncc-1 encodes a cdc2-related kinase required for M phase in meiotic and mitotic cell divisions, but not for S phase. Development 126, 2227–2239 (1999).

    CAS  Google Scholar 

  32. 32

    Stebbins, C. E., Kaelin, W. G. & Pavletich, N. P. Structure of the VHL-ElonginC-ElonginB complex: implications for VHL tumor suppressor function. Science 284, 455–461 (1999).

    CAS  Article  Google Scholar 

  33. 33

    Kamura, T. et al. The elongin BC complex interacts with the conserved SOCS-box motif present in members of the SOCS, ras, WD-40 repeat, and ankyrin repeat families. Genes Dev. 12, 3872– 3881 (1998).

    CAS  Article  Google Scholar 

  34. 34

    Zhang, J.-G. et al. The conserved SOCS box motif in suppressors of cytokine signaling binds to elongins B and C and may couple bound proteins to proteasomal degradation. Proc. Natl Acad. Sci. USA 96, 2071– 2076 (1999).

    CAS  Article  Google Scholar 

  35. 35

    Pause, A., Lee, S., Lonergan, K. M. & Klausner, R. D. The von Hippel-Lindau tumor suppressor gene is required for cell cycle exit upon serum withdrawl. Proc. Natl Acad. Sci. USA 95, 993– 998 (1998).

    CAS  Article  Google Scholar 

  36. 36

    Kim, M. et al. Recombinant adenovirus expressing Von Hippel-Lindau-mediated cell cycle arrest is associated with the induction of cyclin-dependent kinase inhibitor p27Kip1. Biochem. Biophys. Res. Comm. 253, 672–677 (1998).

    CAS  Article  Google Scholar 

  37. 37

    Carrano, A. C., Eytan, E., Hershko, A. & Pagano, M. SKP2 is required for ubiquitin-mediated degradation of the CDK inhibitor p27. Nature Cell Biol. 1, 193–199 (1999).

    CAS  Article  Google Scholar 

  38. 38

    Sutterluty, H. et al. p45SKP2 promotes p27Kip1 degradation and induces S phase in quiescent cells. Nature Cell Biol. 1, 207–214 (1999).

    CAS  Article  Google Scholar 

  39. 39

    Tsvetkov, L. M., Yeh, K.-H., Lee, S.-J., Sun, H. & Zhang, H. p27Kip1 ubiquitination and degradation is regulated by the SCFSkp2 complex through phosphorylated Thr187 in p27. Curr. Biol. 9, 661–664 (1999).

    CAS  Article  Google Scholar 

  40. 40

    Yu, Z.-K., Gervais, J. L. M. & Zhang, H. Human CUL-1 associates with the SKP1/SKP2 complex and regulates p21CIP1/WAF1 and cyclin D proteins. Proc. Natl Acad. Sci. USA 95, 11324–11329 (1998).

    CAS  Article  Google Scholar 

  41. 41

    Kirby, C., Kusch, M. & Kemphues, K. Mutation in the par genes of Caenorhabditis elegans affect cytoplasmic reorganization during the first cell cycle. Dev. Biol. 142, 203–215 (1990).

    CAS  Article  Google Scholar 

  42. 42

    Plasterk, R. H. A. in Caenorhabditis elegans: Modern Biological Analysis of an Organism. Methods in Cell Biology Vol. 48 (eds Epstein, H. F. & Shakes, D. C.) 59–80 (Academic, San Diego, 1995).

    Google Scholar 

  43. 43

    The C. elegans Sequencing Consortium. Genome sequence of the nematode C. elegans: a platform for investigating biology. Science 282, 2012–2018 (1998).

  44. 44

    Krause, M. & Hirsh, D. A trans-spliced leader on actin mRNA in C. elegans. Cell 49, 753– 761 (1987).

    CAS  Article  Google Scholar 

  45. 45

    Thompson, J. D., Higgins, D. G. & Gibson, T. J. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22, 4673–4680 (1994).

    CAS  Article  Google Scholar 

  46. 46

    Swofford, D. L. PAUP: Phylogenetic Analysis Using Parsimony Version 3.1. (Illinois Nat. Hist. Survey, Champaign, IL, 1993).

  47. 47

    Harlow, E. & Lane, D. Antibodies (A Laboratory Manual) (Cold Spring Harb. Lab., Cold Spring Harb., NY, 1988).

    Google Scholar 

  48. 48

    Miller, D. M. & Shakes, D. C. in Caenorhabditis elegans: Modern Biological Analysis of an Organism. Methods in Cell Biology Vol. 48 (eds Epstein, H. F. & Shakes, D. C.) 365–394 (Academic, San Diego, 1995).

    Google Scholar 

  49. 49

    Schumacher, J. M., Ashcroft, N., Donovan, P. J. & Golden, A. A highly conserved centrosomal kinase, AIR-1, is required for accurate cell cycle progression and segregation of developmental factors in Caenorhabditis elegans embryos. Development 125, 4391–4402 (1998).

    CAS  Google Scholar 

  50. 50

    Seydoux, G. & Fire, A. in Caenorhabditis elegans: Modern Biological Analysis of an Organism. Methods in Cell Biology Vol. 48 (eds Epstein, H. F. & Shakes, D. C.) 323–339 (Academic, San Diego, 1995).

    Google Scholar 

  51. 51

    Johnson, C. at 11th International C. elegans meeting May 28–June 1, 1997 (Univ. Wisconsin, Madison, WI, USA).

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Acknowledgements

We thank the Caenorhabditis Genetics Center for C. elegans strains; Y. Kohara for cDNA clones; A. Golden for anti-AIR-1 antibody; R.J. Barstead for a cDNA library; M. Farmer, M. Fechheimer, P. Shen and H. Cai for technical advice; V. Ambros and J. Rothman for communicating results before publication; the Genome Sequencing Consortium for C. elegans genomic sequence and cosmids; T. Schedl for helpful discussions; C. Johnson and L. Liu for providing a second cul-2 deletion allele, which produced an identical phenotype to that produced by allele ek1; and M. Pagano, G. Seydoux and C. Norris for critical reading of the manuscript. This project was supported by NIH grant R01 GM55297 and HFSPO grant RG-229/98 (to E.T.K.).

Correspondence and requests for materials should be addressed to E.T.K. The cki-1 and cki-2 cDNA sequences have been deposited at GenBank under accession numbers AF179358 and AF179359, respectively.

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Correspondence to Edward T. Kipreos.

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Feng, H., Zhong, W., Punkosdy, G. et al. CUL-2 is required for the G1-to-S-phase transition and mitotic chromosome condensation in Caenorhabditis elegans. Nat Cell Biol 1, 486–492 (1999). https://doi.org/10.1038/70272

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