Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

A new family of RhoGEFs activates the Rop molecular switch in plants

Abstract

In plants, the small GTP-binding proteins called Rops work as signalling switches that control growth, development and plant responses to various environmental stimuli1,2,3. Rop proteins (Rho of plants, Rac-like and AtRac in Arabidopsis thaliana) belong to the Rho family of Ras-related GTP-binding proteins that turn on signalling pathways by switching from a GDP-bound inactive to a GTP-bound active conformation4,5. Activation depends on guanine nucleotide exchange factors (GEFs) that catalyse the otherwise slow GDP dissociation for subsequent GTP binding6. Although numerous RhoGEFs exist in animals and yeasts7, no Rop-specific GEFs have yet been identified in plants and so Rop activation has remained elusive1,2,3,8. Here we describe a new family of RhoGEF proteins that are exclusive to plants. We define a unique domain within these RopGEFs, termed PRONE (plant-specific Rop nucleotide exchanger), which is exclusively active towards members of the Rop subfamily. It increases nucleotide dissociation from Rop more than a thousand-fold and forms a tight complex with nucleotide-free Rop. RopGEFs may represent the missing link in signal transduction from receptor kinases to Rops and their identification has implications for the evolution of the Rho molecular switch.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Figure 1: Identification of RopGEFs from Arabidopsis thaliana.
Figure 2: The plant RopGEF family.
Figure 3: Functional characterization of PRONE as genuine GEF domain.
Figure 4: RopGEF1 expression and specificity for Rops.

References

  1. Zheng, Z. & Yang, Z. The Rop GTPase: an emerging signalling switch in plants. Plant Mol. Biol. 44, 1–9 (2000)

    CAS  Article  Google Scholar 

  2. Valster, A. H., Hepler, P. K. & Chernoff, J. Plant GTPases: the Rhos in bloom. Trends Cell Biol. 10, 141–146 (2000)

    CAS  Article  Google Scholar 

  3. Yang, Z. Small GTPases: versatile signalling switches in plants. Plant Cell, S375–S388 (2002)

  4. Bourne, H. R., Sanders, D. A. & McCormick, F. The GTPase superfamily: a conserved switch for diverse cell functions. Nature 348, 125–132 (1990)

    ADS  CAS  Article  Google Scholar 

  5. Bourne, H. R., Sanders, D. A. & McCormick, F. The GTPase superfamily: conserved structure and molecular mechanism. Nature 349, 117–126 (1991)

    ADS  CAS  Article  Google Scholar 

  6. Cherfils, J. & Chardin, P. GEFs: structural basis for their activation of small GTP-binding proteins. Trends Biochem. Sci. 24, 306–311 (1999)

    CAS  Article  Google Scholar 

  7. Rossman, K. L., Der, C. J. & Sondek, J. GEF means go: turning on the Rho GTPases with guanine nucleotide-exchange factors. Nature Rev. Mol. Cell Biol. 6, 167–180 (2005)

    CAS  Article  Google Scholar 

  8. Vernoud, V., Horton, A. C., Yang, Z. & Nielsen, E. Analysis of the small GTPase gene superfamily of Arabidopsis . Plant Physiol. 131, 1191–1208 (2003)

    CAS  Article  Google Scholar 

  9. Hardt, W. D., Chen, L. M., Schuebel, K. E., Bustelo, X. R. & Galán, J. E. S. typhimurium encodes an activator of Rho GTPases that induces membrane ruffling and nuclear reponses in the host cell. Cell 93, 815–826 (1998)

    CAS  Article  Google Scholar 

  10. Rudolph, M. G. et al. Biochemical analysis of SopE from Salmonella typhimurium, a highly efficient guanosine nucleotide exchange factor for RhoGTPases. J. Biol. Chem. 274, 30501–30509 (1999)

    CAS  Article  Google Scholar 

  11. Brugnera, E. et al. Unconventional Rac-GEF activity is mediated through the Dock180-ELMO complex. Nature Cell Biol. 4, 574–582 (2002)

    CAS  Article  Google Scholar 

  12. Meller, N., Irani-Tehrani, M., Kiosses, W. B., Del Pozo, M. A. & Schwartz, M. A. Zizimin1, a novel Cdc42 activator, reveals a new GEF domain for Rho proteins. Nature Cell Biol. 4, 639–647 (2002)

    CAS  Article  Google Scholar 

  13. Qiu, J. L., Jilk, R., Marks, M. D. & Szymanski, D. B. The Arabidopsis SPIKE1 gene is required for normal cell shape and tissue development. Plant Cell 14, 101–118 (2002)

    CAS  Article  Google Scholar 

  14. Schmidt, G. et al. Biochemical and biological consequences of changing the specificity of p21(ras) from guanosine to xanthosine nucleotides. Oncogene 12, 87–96 (1996)

    CAS  PubMed  Google Scholar 

  15. Cool, R. H. et al. The Ras mutant D119N is both dominant negative and active. Mol. Cell. Biol. 19, 6297–6305 (1999)

    CAS  Article  Google Scholar 

  16. Fan, H. Y., Hu, Y., Tudor, M. & Ma, H. Specific interactions between the K domains of AG and AGLs, members of the MADS domain family of DNA binding proteins. Plant J. 12, 999–1010 (1997)

    CAS  Article  Google Scholar 

  17. Bateman, A. et al. The Pfam protein families database. Nucleic Acids Res. 32, D138–D141 (2004)

    CAS  Article  Google Scholar 

  18. Haeusler, L. C., Blumenstein, L., Stege, P., Dvorsky, R. & Ahmadian, M. R. Comparative functional analysis of the Rac GTPases. FEBS Lett. 55, 556–560 (2003)

    Article  Google Scholar 

  19. Worthylake, D. K., Rossman, K. L. & Sondek, J. Crystal structure of Rac1 in complex with the guanine nucleotide exchange region on Tiam1. Nature 408, 682–688 (2000)

    ADS  CAS  Article  Google Scholar 

  20. Karnoub, A. E. et al. Molecular basis for Rac1 recognition by guanine nucleotide exchange factors. Nature Struct. Biol. 8, 1037–1041 (2001)

    CAS  Article  Google Scholar 

  21. Robbe, K., Otto-Bruc, A., Chardin, P. & Antonny, B. Dissociation of GDP dissociation inhibitor and membrane translocation are required for efficient activation of Rac by the Dbl homology-pleckstrin homology region of Tiam. J. Biol. Chem. 278, 4756–4762 (2003)

    CAS  Article  Google Scholar 

  22. Kaothien, P. et al. Kinase partner protein interacts with the LePRK1 and LePRK2 receptor kinases and plays a role in polarized pollen tube growth. Plant J. 42, 492–503 (2005)

    CAS  Article  Google Scholar 

  23. Baldauf, S. L. The deep roots of eukaryotes. Science 300, 1703–1706 (2003)

    ADS  CAS  Article  Google Scholar 

  24. Ho, S. N., Hunt, H. D., Horton, R. M., Pullen, J. K. & Pease, L. R. Site-directed mutagenesis by overlap-extension using the polymerase chain reaction. Gene 77, 51–59 (1989)

    CAS  Article  Google Scholar 

  25. Harper, J. W., Adami, G. R., Wei, N., Keyomarsi, K. & Elledge, S. J. The p21 Cdk-interacting protein Cip1 is a potent inhibitor of G1 cyclin-dependent kinases. Cell 75, 805–816 (1993)

    CAS  Article  Google Scholar 

  26. Lenzen, C., Cool, R. H., Prinz, H., Kuhlmann, J. & Wittinghofer, A. Kinetic analysis by fluorescence of the interaction between Ras and the catalytic domain of the guanine nucleotide exchange factor Cdc25Mm. Biochemistry 37, 7420–7430 (1998)

    CAS  Article  Google Scholar 

  27. John, J. et al. Kinetics of interaction of nucleotides with nucleotide-free H-ras p21. Biochemistry 29, 6058–6065 (1990)

    CAS  Article  Google Scholar 

  28. Laemmli, U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685 (1970)

    ADS  CAS  Article  Google Scholar 

  29. Lenzen, C., Cool, R. H. & Wittinghofer, A. Analysis of intrinsic and Cdc25-stimulated guanine nucleotide exchange of p21ras-nucleotide complexes by fluorescence measurements. Methods Enzymol. 255, 95–109 (1995)

    CAS  Article  Google Scholar 

  30. Bustin, S. A. Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems. J. Mol. Endocrinol. 29, 23–39 (2002)

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank C. Koerner, T. Mitroshina and D. Kuehlmann for technical assistance; R. Ahmadian for purified Rac1, Cdc42, RhoA, Anti-GST antibodies and for helpful discussions; The Arabidopsis Biological Resource Center for the two-hybrid cDNA library (CD4-30) and pAS2; E. W. Weiler for plant material; K. Unfried and J. Abel for providing the LightCycler, and K. Bierhals for critical reading of the manuscript. This work was supported by the Deutsche Forschungsgemeinschaft. C.T. thanks the Boehringer Ingelheim Fonds for support.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Antje Berken.

Ethics declarations

Competing interests

Accession numbers for the plant RopGEF proteins in Arabidopsis thaliana, Oryza sativa, Medicago truncatula and Lycopersicon esculentum are listed in Fig. 2. Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Figure S1

This figure displays a western blot showing the expression of the bait protein Rop4(D121N) and the prey proteins RopGEF1, RopGEF2 and SpkDHR2 in yeast cells that were tested for two-hybrid interactions. (PDF 85 kb)

Supplementary Figure S2

This figure shows a multiple sequence alignment of RopGEF representatives from different plant species and highlights the central part composed of three highly conserved regions C1, C2 and C3. (PDF 1871 kb)

Supplementary Figure Legends

Full text descriptions to accompany the above Supplementary Figures. (DOC 24 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Berken, A., Thomas, C. & Wittinghofer, A. A new family of RhoGEFs activates the Rop molecular switch in plants. Nature 436, 1176–1180 (2005). https://doi.org/10.1038/nature03883

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature03883

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing