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.

  • Article
  • Published:

Gene expression signatures and small-molecule compounds link a protein kinase to Plasmodium falciparum motility

Nature Chemical Biology volume 4, pages 347–356 (2008)Cite this article

Abstract

Calcium-dependent protein kinases play a crucial role in intracellular calcium signaling in plants, some algae and protozoa. In Plasmodium falciparum, calcium-dependent protein kinase 1 (PfCDPK1) is expressed during schizogony in the erythrocytic stage as well as in the sporozoite stage. It is coexpressed with genes that encode the parasite motor complex, a cellular component required for parasite invasion of host cells, parasite motility and potentially cytokinesis. A targeted gene-disruption approach demonstrated that pfcdpk1 seems to be essential for parasite viability. An in vitro biochemical screen using recombinant PfCDPK1 against a library of 20,000 compounds resulted in the identification of a series of structurally related 2,6,9-trisubstituted purines. Compound treatment caused sudden developmental arrest at the late schizont stage in P. falciparum and a large reduction in intracellular parasites in Toxoplasma gondii, which suggests a possible role for PfCDPK1 in regulation of parasite motility during egress and invasion.

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

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: pfcdpk1 is coexpressed with genes involved in parasite motility.
Figure 2: Comparative genome hybridization for knockout strains.
Figure 3: Effect of purfalcamine on parasitemia.
Figure 4: Representative images from a T. gondii invasion assay carried out in the presence of varying concentrations of purfalcamine.
Figure 5: Structure-based ligand docking analysis.

Similar content being viewed by others

References

  1. Snow, R.W., Guerra, C.A., Noor, A.M., Myint, H.Y. & Hay, S.I. The global distribution of clinical episodes of Plasmodium falciparum malaria. Nature 434, 214–217 (2005).

    Article  CAS  Google Scholar 

  2. Garcia, C.R. Calcium homeostasis and signaling in the blood-stage malaria parasite. Parasitol. Today 15, 488–491 (1999).

    Article  CAS  Google Scholar 

  3. Gazarini, M.L., Thomas, A.P., Pozzan, T. & Garcia, C.R. Calcium signaling in a low calcium environment: how the intracellular malaria parasite solves the problem. J. Cell Biol. 161, 103–110 (2003).

    Article  CAS  Google Scholar 

  4. Nagamune, K. & Sibley, L.D. Comparative genomic and phylogenetic analyses of calcium ATPases and calcium-regulated proteins in the apicomplexa. Mol. Biol. Evol. 23, 1613–1627 (2006).

    Article  CAS  Google Scholar 

  5. Ward, P., Equinet, L., Packer, J. & Doerig, C. Protein kinases of the human malaria parasite Plasmodium falciparum: the kinome of a divergent eukaryote. BMC Genomics 5, 79 (2004).

    Article  Google Scholar 

  6. Billker, O. et al. Calcium and a calcium-dependent protein kinase regulate gamete formation and mosquito transmission in a malaria parasite. Cell 117, 503–514 (2004).

    Article  CAS  Google Scholar 

  7. Ishino, T., Orito, Y., Chinzei, Y. & Yuda, M. A calcium-dependent protein kinase regulates Plasmodium ookinete access to the midgut epithelial cell. Mol. Microbiol. 59, 1175–1184 (2006).

    Article  CAS  Google Scholar 

  8. Siden-Kiamos, I. et al. Plasmodium berghei calcium-dependent protein kinase 3 is required for ookinete gliding motility and mosquito midgut invasion. Mol. Microbiol. 60, 1355–1363 (2006).

    Article  CAS  Google Scholar 

  9. Le Roch, K.G. et al. Discovery of gene function by expression profiling of the malaria parasite life cycle. Science 301, 1503–1508 (2003).

    Article  CAS  Google Scholar 

  10. Moskes, C. et al. Export of Plasmodium falciparum calcium-dependent protein kinase 1 to the parasitophorous vacuole is dependent on three N-terminal membrane anchor motifs. Mol. Microbiol. 54, 676–691 (2004).

    Article  Google Scholar 

  11. Sanders, P.R. et al. Distinct protein classes including novel merozoite surface antigens in Raft-like membranes of Plasmodium falciparum. J. Biol. Chem. 280, 40169–40176 (2005).

    Article  CAS  Google Scholar 

  12. Young, J.A. et al. The Plasmodium falciparum sexual development transcriptome: a microarray analysis using ontology-based pattern identification. Mol. Biochem. Parasitol. 143, 67–79 (2005).

    Article  CAS  Google Scholar 

  13. Zhou, Y. et al. Evidence-based annotation of the malaria parasite's genome using comparative expression profiling. PLoS ONE 3, e1570 (2008).

    Article  Google Scholar 

  14. Baum, J. et al. A conserved molecular motor drives cell invasion and gliding motility across malaria life cycle stages and other apicomplexan parasites. J. Biol. Chem. 281, 5197–5208 (2006).

    Article  CAS  Google Scholar 

  15. Heintzelman, M.B. Cellular and molecular mechanics of gliding locomotion in eukaryotes. Int. Rev. Cytol. 251, 79–129 (2006).

    Article  CAS  Google Scholar 

  16. Lovett, J.L. & Sibley, L.D. Intracellular calcium stores in Toxoplasma gondii govern invasion of host cells. J. Cell Sci. 116, 3009–3016 (2003).

    Article  CAS  Google Scholar 

  17. Bergman, L.W. et al. Myosin A tail domain interacting protein (MTIP) localizes to the inner membrane complex of Plasmodium sporozoites. J. Cell Sci. 116, 39–49 (2003).

    Article  CAS  Google Scholar 

  18. Green, J.L. et al. The MTIP-myosin A complex in blood stage malaria parasites. J. Mol. Biol. 355, 933–941 (2006).

    Article  CAS  Google Scholar 

  19. Meissner, M., Schluter, D. & Soldati, D. Role of Toxoplasma gondii myosin A in powering parasite gliding and host cell invasion. Science 298, 837–840 (2002).

    Article  CAS  Google Scholar 

  20. Carruthers, V.B. & Sibley, L.D. Mobilization of intracellular calcium stimulates microneme discharge in Toxoplasma gondii. Mol. Microbiol. 31, 421–428 (1999).

    Article  CAS  Google Scholar 

  21. Duraisingh, M.T., Triglia, T. & Cowman, A.F. Negative selection of Plasmodium falciparum reveals targeted gene deletion by double crossover recombination. Int. J. Parasitol. 32, 81–89 (2002).

    Article  CAS  Google Scholar 

  22. Zhang, J.H., Chung, T.D. & Oldenburg, K.R. A simple statistical parameter for use in evaluation and validation of high throughput screening assays. J. Biomol. Screen. 4, 67–73 (1999).

    Article  CAS  Google Scholar 

  23. Ding, S., Gray, N.S., Wu, X., Ding, Q. & Schultz, P.G. A combinatorial scaffold approach toward kinase-directed heterocycle libraries. J. Am. Chem. Soc. 124, 1594–1596 (2002).

    Article  CAS  Google Scholar 

  24. Bennett, T.N. et al. Novel, rapid, and inexpensive cell-based quantification of antimalarial drug efficacy. Antimicrob. Agents Chemother. 48, 1807–1810 (2004).

    Article  CAS  Google Scholar 

  25. Smilkstein, M., Sriwilaijaroen, N., Kelly, J.X., Wilairat, P. & Riscoe, M. Simple and inexpensive fluorescence-based technique for high-throughput antimalarial drug screening. Antimicrob. Agents Chemother. 48, 1803–1806 (2004).

    Article  CAS  Google Scholar 

  26. Chulay, J.D., Haynes, J.D. & Diggs, C.L. Plasmodium falciparum: assessment of in vitro growth by [3H]hypoxanthine incorporation. Exp. Parasitol. 55, 138–146 (1983).

    Article  CAS  Google Scholar 

  27. Duraisingh, M.T. et al. Linkage disequilibrium between two chromosomally distinct loci associated with increased resistance to chloroquine in Plasmodium falciparum. Parasitology 121, 1–7 (2000).

    Article  CAS  Google Scholar 

  28. Fotie, J. et al. Lupeol long-chain fatty acid esters with antimalarial activity from Holarrhena floribunda. J. Nat. Prod. 69, 62–67 (2006).

    Article  CAS  Google Scholar 

  29. Florens, L. et al. A proteomic view of the Plasmodium falciparum life cycle. Nature 419, 520–526 (2002).

    Article  CAS  Google Scholar 

  30. Baldwin, J. et al. High-throughput screening for potent and selective inhibitors of Plasmodium falciparum dihydroorotate dehydrogenase. J. Biol. Chem. 280, 21847–21853 (2005).

    Article  CAS  Google Scholar 

  31. Barrera, L., Benner, C., Tao, Y.C., Winzeler, E. & Zhou, Y. Leveraging two-way probe-level block design for identifying differential gene expression with high-density oligonucleotide arrays. BMC Bioinformatics 5, 42 (2004).

    Article  Google Scholar 

  32. Oshiro, G. et al. Parallel identification of new genes in Saccharomyces cerevisiae. Genome Res. 12, 1210–1220 (2002).

    Article  CAS  Google Scholar 

  33. Ashburner, M. et al. Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat. Genet. 25, 25–29 (2000).

    Article  CAS  Google Scholar 

  34. Suthram, S., Sittler, T. & Ideker, T. The Plasmodium protein network diverges from those of other eukaryotes. Nature 438, 108–112 (2005).

    Article  CAS  Google Scholar 

  35. Bosch, J. et al. The closed MTIP-myosin A-tail complex from the malaria parasite invasion machinery. J. Mol. Biol. 372, 77–88 (2007).

    Article  CAS  Google Scholar 

  36. Kidgell, C. et al. A systematic map of genetic variation in Plasmodium falciparum. PLoS Pathog. 2, e57 (2006).

    Article  Google Scholar 

  37. Peters, W. & Robinson, B.L. in Handbook of Animal Models of Infection (eds. Zak, O. & Sande, M.) 757–773 (Academic, London, 1999).

    Book  Google Scholar 

  38. Srinivasan, A.N. & Krupa, A. A genomic perspective of protein kinases in Plasmodium falciparum. Proteins 58, 180–189 (2005).

    CAS  PubMed  Google Scholar 

  39. Wetzel, D.M., Chen, L.A., Ruiz, F.A., Moreno, S.N. & Sibley, L.D. Calcium-mediated protein secretion potentiates motility in Toxoplasma gondii. J. Cell Sci. 117, 5739–5748 (2004).

    Article  CAS  Google Scholar 

  40. Keeley, A. & Soldati, D. The glideosome: a molecular machine powering motility and host-cell invasion by Apicomplexa. Trends Cell Biol. 14, 528–532 (2004).

    Article  CAS  Google Scholar 

  41. Gantt, S. et al. Antibodies against thrombospondin-related anonymous protein do not inhibit Plasmodium sporozoite infectivity in vivo. Infect. Immun. 68, 3667–3673 (2000).

    Article  CAS  Google Scholar 

  42. Kieschnick, H., Wakefield, T., Narducci, C.A. & Beckers, C. Toxoplasma gondii attachment to host cells is regulated by a calmodulin-like domain protein kinase. J. Biol. Chem. 276, 12369–12377 (2001).

    Article  CAS  Google Scholar 

  43. Karcher, R.L. et al. Cell cycle regulation of myosin-V by calcium/calmodulin-dependent protein kinase II. Science 293, 1317–1320 (2001).

    Article  CAS  Google Scholar 

  44. Kugelstadt, D., Winter, D., Pluckhahn, K., Lehmann, W.D. & Kappes, B. Raf kinase inhibitor protein affects activity of Plasmodium falciparum calcium-dependent protein kinase 1. Mol. Biochem. Parasitol. 151, 111–117 (2007).

    Article  CAS  Google Scholar 

  45. LaCount, D.J. et al. A protein interaction network of the malaria parasite Plasmodium falciparum. Nature 438, 103–107 (2005).

    Article  CAS  Google Scholar 

  46. Cardone, M.H. et al. Regulation of cell death protease caspase-9 by phosphorylation. Science 282, 1318–1321 (1998).

    Article  CAS  Google Scholar 

  47. Arastu-Kapur, S. et al. Identification of proteases that regulate erythrocyte rupture by the malaria parasite Plasmodium falciparum. Nat. Chem. Biol. 4, 203–213 (2008).

    Article  CAS  Google Scholar 

  48. Yeoh, S. et al. Subcellular discharge of a serine protease mediates release of invasive malaria parasites from host erythrocytes. Cell 131, 1072–1083 (2007).

    Article  CAS  Google Scholar 

  49. Lambros, C. & Vanderberg, J.P. Synchronization of Plasmodium falciparum erythrocytic stages in culture. J. Parasitol. 65, 418–420 (1979).

    Article  CAS  Google Scholar 

  50. Carey, K.L., Westwood, N.J., Mitchison, T.J. & Ward, G.E. A small-molecule approach to studying invasive mechanisms of Toxoplasma gondii. Proc. Natl. Acad. Sci. USA 101, 7433–7438 (2004).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank V. Nussenzweig (New York University) for providing oocyst and sporozoite stages of P. yoelii and A. Godfrey-Certner for her assistance with the T. gondii invasion assays. This work was supported by a grant to E.A.W. from the Ellison Foundation, the Keck Foundation and the US National Institutes of Health (AI059472-02), a grant to G.E.W. from the US National Institutes of Health (AI054961), and grants to the Genomics Institute of the Novartis Research Foundation from the Wellcome Trust and the Medicines for Malaria Venture.

Author information

Authors and Affiliations

  1. Department of Cell Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, ICND202, La Jolla, 92037, California, USA

    Nobutaka Kato, Ghislain Breton, Kerstin Henson, Jeffrey Johnson, Felix Marr, Vandana Ramachandran, Peter G Schultz & Elizabeth A Winzeler

  2. Genomics Institute of the Novartis Research Foundation, 10675 John Jay Hopkins Drive, San Diego, 92121, California, USA

    Nobutaka Kato, Tomoyo Sakata, Karine G Le Roch, Advait Nagle, Carsten Andersen, Badry Bursulaya, Kerstin Henson, Daniel Mason, Case McNamara, David Plouffe, Muriel Spooner, Tove Tuntland, Yingyao Zhou, Eric C Peters, Arnab Chatterjee, Peter G Schultz, Nathanael Gray & Elizabeth A Winzeler

  3. Michael Heidelberger Division of Immunology, Department of Pathology, New York University School of Medicine, 550 First Avenue, New York, 10016, New York, USA

    Kota Arun Kumar

  4. Department of Microbiology and Molecular Genetics, University of Vermont, 95 Carrigan Drive, Burlington, 05405, Vermont, USA

    Gary E Ward

  5. Biochemistry Department, University of Nevada, 1664 North Virginia Street, Reno, 89557, Nevada, USA

    Jeffrey Harper

Authors
  1. Nobutaka Kato
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tomoyo Sakata
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ghislain Breton
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Karine G Le Roch
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Advait Nagle
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Carsten Andersen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Badry Bursulaya
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Kerstin Henson
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jeffrey Johnson
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Kota Arun Kumar
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Felix Marr
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Daniel Mason
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. Case McNamara
    View author publications

    You can also search for this author in PubMed Google Scholar

  14. David Plouffe
    View author publications

    You can also search for this author in PubMed Google Scholar

  15. Vandana Ramachandran
    View author publications

    You can also search for this author in PubMed Google Scholar

  16. Muriel Spooner
    View author publications

    You can also search for this author in PubMed Google Scholar

  17. Tove Tuntland
    View author publications

    You can also search for this author in PubMed Google Scholar

  18. Yingyao Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  19. Eric C Peters
    View author publications

    You can also search for this author in PubMed Google Scholar

  20. Arnab Chatterjee
    View author publications

    You can also search for this author in PubMed Google Scholar

  21. Peter G Schultz
    View author publications

    You can also search for this author in PubMed Google Scholar

  22. Gary E Ward
    View author publications

    You can also search for this author in PubMed Google Scholar

  23. Nathanael Gray
    View author publications

    You can also search for this author in PubMed Google Scholar

  24. Jeffrey Harper
    View author publications

    You can also search for this author in PubMed Google Scholar

  25. Elizabeth A Winzeler
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Elizabeth A Winzeler.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–8, Supplementary Tables 1 and 2, and Supplementary Methods (PDF 3468 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Kato, N., Sakata, T., Breton, G. et al. Gene expression signatures and small-molecule compounds link a protein kinase to Plasmodium falciparum motility. Nat Chem Biol 4, 347–356 (2008). https://doi.org/10.1038/nchembio.87

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nchembio.87

This article is cited by

Search

Advanced 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