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PAK4 kinase activity and somatic mutation promote carcinoma cell motility and influence inhibitor sensitivity

Oncogene volume 32, pages 2114–2120 (2013)Cite this article

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Abstract

Hepatocyte growth factor (HGF) and its receptor (c-Met) are associated with cancer cell motility and invasiveness. p21-activated kinase 4 (PAK4), a potential therapeutic target, is recruited to and activated by c-Met. In response, PAK4 phosphorylates LIM kinase 1 (LIMK1) in an HGF-dependent manner in metastatic prostate carcinoma cells. PAK4 overexpression is known to induce increased cell migration speed but the requirement for kinase activity has not been established. We have used a panel of PAK4 truncations and mutations in a combination of overexpression and RNAi rescue experiments to determine the requirement for PAK4 kinase activity during carcinoma cell motility downstream of HGF. We find that neither the kinase domain alone nor a PAK4 mutant unable to bind Cdc42 is able to fully rescue cell motility in a PAK4-deficient background. Nevertheless, we find that PAK4 kinase activity and associated LIMK1 activity are essential for carcinoma cell motility, highlighting PAK4 as a potential anti-metastatic therapeutic target. We also show here that overexpression of PAK4 harbouring a somatic mutation, E329K, increased the HGF-driven motility of metastatic prostate carcinoma cells. E329 lies within the glycine-rich loop region of the kinase. Our data suggest that E329K mutation leads to a modest increase in kinase activity, conferring resistance to competitive ATP inhibitors in addition to promoting cell migration. The existence of such a mutation may have implications for the development of PAK4-specific competitive ATP inhibitors should PAK4 be further explored for clinical inhibition.

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References

  1. Cecchi F, Rabe DC, Bottaro DP . Targeting the HGF/Met signalling pathway in cancer. Eur J Cancer 2010; 46: 1260–1270.

    Article  CAS  Google Scholar 

  2. Knudsen BS, Woude GV . Showering c-MET-dependent cancers with drugs. Curr opin Genetics Dev 2008; 18: 87–96.

    Article  CAS  Google Scholar 

  3. Lai AZ, Abella JV, Park M . Crosstalk in Met receptor oncogenesis. Trends Cell Biol 2009; 19: 542–551.

    Article  CAS  Google Scholar 

  4. Paliouras GN, Naujokas MA, Park M . Pak4, a novel Gab1 binding partner, modulates cell migration and invasion by the Met receptor. Mol Cell Biol 2009; 29: 3018–3032.

    Article  CAS  Google Scholar 

  5. Abo A, Qu J, Cammarano MS, Dan C, Fritsch A, Baud V et al. PAK4, a novel effector for Cdc42Hs, is implicated in the reorganization of the actin cytoskeleton and in the formation of filopodia. EMBO J 1998; 17: 6527–6540.

    Article  CAS  Google Scholar 

  6. Callow MG, Zozulya S, Gishizky ML, Jallal B, Smeal T . PAK4 mediates morphological changes through the regulation of GEF-H1. J Cell Sci 2005; 118: 1861–1872.

    Article  CAS  Google Scholar 

  7. Wells CM, Whale AD, Parsons M, Masters JR, Jones GE . PAK4: a pluripotent kinase that regulates prostate cancer cell adhesion. J Cell Sci 2010; 123: 1663–1673.

    Article  Google Scholar 

  8. Li Z, Zhang H, Lundin L, Thullberg M, Liu Y, Wang Y et al. p21-activated kinase 4 phosphorylation of integrin β5 Ser-759 and Ser-762 regulates cell migration. J Biol Chem 2010; 285: 23699–23710.

    Article  CAS  Google Scholar 

  9. Dan C, Kelly A, Bernard O, Minden A . Cytoskeletal changes regulated by the PAK4 serine/threonine kinase are mediated by LIM kinase 1 and cofilin. J Biol Chem 2001; 276: 32115–32121.

    Article  CAS  Google Scholar 

  10. Soosairajah J, Maiti S, Wiggan O, Sarmiere P, Moussi N, Sarcevic B et al. Interplay between components of a novel LIM kinase-slingshot phosphatase complex regulates cofilin. EMBO J 2005; 24: 473–486.

    Article  CAS  Google Scholar 

  11. Lai FPL, Szczodrak M, Block J, Faix J, Breitsprecher D, Mannherz HG et al. Arp2/3 complex interactions and actin network turnover in lamellipodia. EMBO J 2008; 27: 982–992.

    Article  CAS  Google Scholar 

  12. Ghosh M, Song X, Mouneimne G, Sidani M, Lawrence DS, Condeelis JS et al. Cofilin promotes actin polymerization and defines the direction of cell motility. Science 2004; 304: 743–746.

    Article  CAS  Google Scholar 

  13. Callow MG, Clairvoyant F, Zhu S, Schryver B, Whyte DB, Bischoff JR et al. Requirement for PAK4 in the anchorage-independent growth of human cancer cell lines. J Biol Chem 2002; 277: 550–558.

    Article  CAS  Google Scholar 

  14. Liu Y, Xiao H, Tian Y, Nekrasova T, Hao X, Lee HJ et al. The pak4 protein kinase plays a key role in cell survival and tumorigenesis in athymic mice. Mol Cancer Res 2008; 6: 215–224.

    Google Scholar 

  15. Whale A, Hashim FN, Fram S, Jones GE, Wells CM . Signalling to cancer cell invasion through PAK family kinases. Front Biosci 2011; 16: 849–864.

    Article  CAS  Google Scholar 

  16. Parsons DW, Wang TL, Samuels Y, Bardelli A, Cummins JM, DeLong L et al. Colorectal cancer: mutations in a signalling pathway. Nature 2005; 436: 792.

    Article  CAS  Google Scholar 

  17. Kimmelman AC, Hezel AF, Aguirre AJ, Zheng H, Paik J, Ying H et al. Genomic alterations link Rho family of GTPases to the highly invasive phenotype of pancreas cancer. Proc Natl Acad Sci USA 2008; 105: 19372–19377.

    Article  CAS  Google Scholar 

  18. Li Z, Lock JG, Olofsson H, Kowalewski JM, Teller S, Liu Y et al. Integrin-mediated cell attachment induces a PAK4-dependent feedback loop regulating cell adhesion through modified integrin αvβ5 clustering and turnover. Mol Biol Cell 2010; 21: 3317–3329.

    Article  CAS  Google Scholar 

  19. Siu MKY, Chan HY, Konga DSH, Wong ESY, Wong OGW, Ngan HYS et al. p21-activated kinase 4 regulates ovarian cancer cell proliferation, migration, and invasion and contributes to poor prognosis in patients. Proc Natl Acad Sci USA 2010; 107: 18622–18627.

    Article  CAS  Google Scholar 

  20. Eswaran J, Soundararajan M, Knapp S . Targeting group II PAKs in cancer and metastasis. Cancer Metastasis Rev 2009; 28: 209–217.

    Article  CAS  Google Scholar 

  21. Zhao Z, Manser E . Do PAKs make good drug targets? Faculty of 1000 2010; 2: 70.

    Google Scholar 

  22. Murray BW, Guoa C, Piraino J, Westwick JK, Zhang C, Lamerdin J et al. Small-molecule p21-activated kinase inhibitor PF-3758309 is a potent inhibitor of oncogenic signaling and tumor growth. Proc Natl Acad Sci USA 2010; 107: 9446–9451.

    Article  CAS  Google Scholar 

  23. Finkelstein E, Chang W, Chao P-HG, Gruber D, Minden A, Hung CT et al. Roles of microtubules, cell polarity and adhesion in electric-field-mediated motility of 3T3 fibroblasts. J Cell Sci 2004; 117: 1533–1545.

    Article  CAS  Google Scholar 

  24. Ahmed T, Shea K, Masters JR, Jones GE, Wells CMA . PAK4-LIMK1 pathway drives prostate cancer cell migration downstream of HGF. Cell Signal 2008; 20: 1320–1328.

    Article  CAS  Google Scholar 

  25. Scott RW, Hooper S, Crighton D, Li A, König I, Munro J et al. LIM kinases are required for invasive path generation by tumor and tumor-associated stromal cells. J Cell Biol 2010; 191: 169–185.

    Article  CAS  Google Scholar 

  26. Yoshioka K, Foletta V, Bernard O, Itoh K . A role for LIM kinase in cancer invasion. Proc Natl Acad Sci USA 2003; 100: 7247–7252.

    Article  CAS  Google Scholar 

  27. Baldassa S, Calogero AM, Colombo G, Zippel R, Gnesutta N . N-Terminal interaction domain implicates PAK4 in translational regulation and reveals novel cellular localization signals. J Cell Physiol 2010; 224: 722–733.

    Article  CAS  Google Scholar 

  28. Eswaran J, Lee WH, Debreczeni JE, Filippakopoulos P, Turnbull A, Fedorov O et al. Crystal structures of the p21-activated kinases PAK4, PAK5, and PAK6 reveal catalytic domain plasticity of active group II PAKs. Structure 2007; 15: 201–213.

    Article  CAS  Google Scholar 

  29. Grant BD, Hemmer W, Tsigelny I, Adams JA, Taylor SS . Kinetic analyses of mutations in the glycine-rich loop of cAMP-dependent protein kinase. Biochemistry 1998; 37: 7708–7715.

    Article  CAS  Google Scholar 

  30. Barouch-Bentov R, Che J, Lee CC, Yang Y, Herman A, Jia Y et al. A conserved salt bridge in the G loop of multiple protein kinases is important for catalysis and for in vivo Lyn function. Mol Cell 2009; 33: 43–52.

    Article  CAS  Google Scholar 

  31. Griswold IJ, MacPartlin M, Bumm T, Goss VL, O’Hare T, Lee KA et al. Kinase domain mutants of Bcr-Abl exhibit altered transformation potency, kinase activity, and substrate utilization, irrespective of sensitivity to imatinib. Mol Cell Biol 2006; 26: 6082–6093.

    Article  CAS  Google Scholar 

  32. Yamamoto M, Kurosu T, Kakihana K, Mizuchi D, Miura O . The two major imatinib resistance mutations E255K and T315I enhance the activity of BCR/ABL fusion kinase. Biochem Biophys Res Commun 2004; 319: 1272–1275.

    Article  CAS  Google Scholar 

  33. Skaggs BJ, Gorre ME, Ryvkin A, Burgess MR, Xie Y, Han Y et al. Phosphorylation of the ATP-binding loop directs oncogenicity of drug-resistant BCR-ABL mutants. Proc Natl Acad Sci USA 2006; 103: 19466–19471.

    Article  CAS  Google Scholar 

  34. Shah NP, Nicoll JM, Nagar B, Gorre ME, Paquette RL, Kuriyan J et al. Multiple BCR-ABL kinase domain mutations confer polyclonal resistance to the tyrosine kinase inhibitor imatinib (STI571) in chronic phase and blast crisis chronic myeloid leukemia. Cancer Cell 2002; 2: 117–125.

    Article  CAS  Google Scholar 

  35. Pavlovsky C, Kantarjian H, Cortes JE . First-line therapy for chronic myeloid leukemia: past, present, and future. Am J Hematol 2009; 84: 287–293.

    Article  CAS  Google Scholar 

  36. Girdler F, Sessa F, Patercoli S, Villa F, Musacchio A, Taylor S et al. Molecular basis of drug resistance in aurora kinases. Chem Biol 2008; 15: 552–562.

    Article  CAS  Google Scholar 

  37. Zicha D, Dunn G, Jones G . Analyzing chemotaxis using the Dunn direct-viewing chamber. Methods Mol Biol 1997; 75: 449–57.

    CAS  PubMed  Google Scholar 

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Acknowledgements

ADW and work in the laboratories of GEJ and CMW is supported by a grant from the Cancer Research UK. AD is supported by a grant from Breast Cancer Campaign. We would like to thank Matthias Krause for gateway destination and pGIPZ-control shRNA vectors. We also thank Mary Holdom for helpful discussion and advice with protein structure analysis and Mike Olsen for practical advice with using LIMKi.

Author contributions: ADW, GEJ and CMW planned the experiments. ADW, AD and CMW conducted the experiments. MH performed migration analysis. ADW, GEJ and CMW wrote the paper. CW and GEJ contributed equally to the paper.

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  1. Randall Division of Cell and Molecular Biophysics, King’s College London, London, UK

    A D Whale, M Holt & G E Jones

  2. Division of Cancer Studies, King’s College London, London, UK

    A D Whale, A Dart & C M Wells

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  1. A D Whale
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  2. A Dart
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Correspondence to C M Wells.

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Whale, A., Dart, A., Holt, M. et al. PAK4 kinase activity and somatic mutation promote carcinoma cell motility and influence inhibitor sensitivity. Oncogene 32, 2114–2120 (2013). https://doi.org/10.1038/onc.2012.233

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