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. 2001 Jul 15;357(Pt 2):427–435. doi: 10.1042/0264-6021:3570427

Targeting mutants of PTEN reveal distinct subsets of tumour suppressor functions.

N R Leslie 1, D Bennett 1, A Gray 1, I Pass 1, K Hoang-Xuan 1, C P Downes 1
PMCID: PMC1221969  PMID: 11439092

Abstract

The tumour suppressor protein PTEN (phosphatase and tensin homolog deleted on chromosome 10) is a lipid phosphatase which can antagonize the phosphoinositide 3-kinase (PI 3-kinase) signalling pathway, promoting apoptosis and inhibiting cell-cycle progression and cell motility. We show that very little cellular PTEN is associated with the plasma membrane, but that artificial membrane-targeting of PTEN enhances its inhibition of signalling to protein kinase B (PKB). Evidence for potential targeting of PTEN to the membrane through PDZ domain-mediated protein-protein interactions led us to use a PTEN enzyme with a deletion of the C-terminal PDZ-binding sequence, that retains full phosphatase activity against soluble substrates, and to analyse the efficiency of this mutant in different cellular assays. The extreme C-terminal PDZ-binding sequence was dispensable for the efficient down-regulation of cellular PtdIns(3,4,5)P3 levels and a number of PI 3-kinase-dependent signalling activities, including PKB and p70S6K. However, the PDZ-binding sequence was required for the efficient inhibition of cell spreading. The data show that a PTEN mutation, similar to those found in some tumours, affects some functions of the protein but not others, and implicate the deregulation of PTEN-dependent processes other than PKB activation in the development of some tumours. Significantly, this hypothesis is supported by data showing low levels of PKB phosphorylation in a glioblastoma sample carrying a mutation in the extreme C-terminus of PTEN compared with tumours carrying phosphatase-inactivating mutations of the enzyme. Our data show that deregulation of PKB is not a universal feature of tumours carrying PTEN mutations and implicate other processes that may be deregulated in these tumours.

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Selected References

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  1. Adey N. B., Huang L., Ormonde P. A., Baumgard M. L., Pero R., Byreddy D. V., Tavtigian S. V., Bartel P. L. Threonine phosphorylation of the MMAC1/PTEN PDZ binding domain both inhibits and stimulates PDZ binding. Cancer Res. 2000 Jan 1;60(1):35–37. [PubMed] [Google Scholar]
  2. Alessi D. R., James S. R., Downes C. P., Holmes A. B., Gaffney P. R., Reese C. B., Cohen P. Characterization of a 3-phosphoinositide-dependent protein kinase which phosphorylates and activates protein kinase Balpha. Curr Biol. 1997 Apr 1;7(4):261–269. doi: 10.1016/s0960-9822(06)00122-9. [DOI] [PubMed] [Google Scholar]
  3. Alessi D. R., Kozlowski M. T., Weng Q. P., Morrice N., Avruch J. 3-Phosphoinositide-dependent protein kinase 1 (PDK1) phosphorylates and activates the p70 S6 kinase in vivo and in vitro. Curr Biol. 1998 Jan 15;8(2):69–81. doi: 10.1016/s0960-9822(98)70037-5. [DOI] [PubMed] [Google Scholar]
  4. Ali I. U., Schriml L. M., Dean M. Mutational spectra of PTEN/MMAC1 gene: a tumor suppressor with lipid phosphatase activity. J Natl Cancer Inst. 1999 Nov 17;91(22):1922–1932. doi: 10.1093/jnci/91.22.1922. [DOI] [PubMed] [Google Scholar]
  5. Batty I. H., Downes C. P. The inhibition of phosphoinositide synthesis and muscarinic-receptor-mediated phospholipase C activity by Li+ as secondary, selective, consequences of inositol depletion in 1321N1 cells. Biochem J. 1994 Feb 1;297(Pt 3):529–537. doi: 10.1042/bj2970529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bonneau D., Longy M. Mutations of the human PTEN gene. Hum Mutat. 2000;16(2):109–122. doi: 10.1002/1098-1004(200008)16:2<109::AID-HUMU3>3.0.CO;2-0. [DOI] [PubMed] [Google Scholar]
  7. Cantley L. C., Neel B. G. New insights into tumor suppression: PTEN suppresses tumor formation by restraining the phosphoinositide 3-kinase/AKT pathway. Proc Natl Acad Sci U S A. 1999 Apr 13;96(8):4240–4245. doi: 10.1073/pnas.96.8.4240. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cox D., Tseng C. C., Bjekic G., Greenberg S. A requirement for phosphatidylinositol 3-kinase in pseudopod extension. J Biol Chem. 1999 Jan 15;274(3):1240–1247. doi: 10.1074/jbc.274.3.1240. [DOI] [PubMed] [Google Scholar]
  9. Dahia P. L. PTEN, a unique tumor suppressor gene. Endocr Relat Cancer. 2000 Jun;7(2):115–129. doi: 10.1677/erc.0.0070115. [DOI] [PubMed] [Google Scholar]
  10. Furnari F. B., Huang H. J., Cavenee W. K. The phosphoinositol phosphatase activity of PTEN mediates a serum-sensitive G1 growth arrest in glioma cells. Cancer Res. 1998 Nov 15;58(22):5002–5008. [PubMed] [Google Scholar]
  11. Georgescu M. M., Kirsch K. H., Akagi T., Shishido T., Hanafusa H. The tumor-suppressor activity of PTEN is regulated by its carboxyl-terminal region. Proc Natl Acad Sci U S A. 1999 Aug 31;96(18):10182–10187. doi: 10.1073/pnas.96.18.10182. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Georgescu M. M., Kirsch K. H., Kaloudis P., Yang H., Pavletich N. P., Hanafusa H. Stabilization and productive positioning roles of the C2 domain of PTEN tumor suppressor. Cancer Res. 2000 Dec 15;60(24):7033–7038. [PubMed] [Google Scholar]
  13. Haas-Kogan D., Shalev N., Wong M., Mills G., Yount G., Stokoe D. Protein kinase B (PKB/Akt) activity is elevated in glioblastoma cells due to mutation of the tumor suppressor PTEN/MMAC. Curr Biol. 1998 Oct 22;8(21):1195–1198. doi: 10.1016/s0960-9822(07)00493-9. [DOI] [PubMed] [Google Scholar]
  14. Hawkins P. T., Eguinoa A., Qiu R. G., Stokoe D., Cooke F. T., Walters R., Wennström S., Claesson-Welsh L., Evans T., Symons M. PDGF stimulates an increase in GTP-Rac via activation of phosphoinositide 3-kinase. Curr Biol. 1995 Apr 1;5(4):393–403. doi: 10.1016/s0960-9822(95)00080-7. [DOI] [PubMed] [Google Scholar]
  15. Ignatoski K. M., Maehama T., Markwart S. M., Dixon J. E., Livant D. L., Ethier S. P. ERBB-2 overexpression confers PI 3' kinase-dependent invasion capacity on human mammary epithelial cells. Br J Cancer. 2000 Feb;82(3):666–674. doi: 10.1054/bjoc.1999.0979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kraynov V. S., Chamberlain C., Bokoch G. M., Schwartz M. A., Slabaugh S., Hahn K. M. Localized Rac activation dynamics visualized in living cells. Science. 2000 Oct 13;290(5490):333–337. doi: 10.1126/science.290.5490.333. [DOI] [PubMed] [Google Scholar]
  17. Lachyankar M. B., Sultana N., Schonhoff C. M., Mitra P., Poluha W., Lambert S., Quesenberry P. J., Litofsky N. S., Recht L. D., Nabi R. A role for nuclear PTEN in neuronal differentiation. J Neurosci. 2000 Feb 15;20(4):1404–1413. doi: 10.1523/JNEUROSCI.20-04-01404.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lee J. O., Yang H., Georgescu M. M., Di Cristofano A., Maehama T., Shi Y., Dixon J. E., Pandolfi P., Pavletich N. P. Crystal structure of the PTEN tumor suppressor: implications for its phosphoinositide phosphatase activity and membrane association. Cell. 1999 Oct 29;99(3):323–334. doi: 10.1016/s0092-8674(00)81663-3. [DOI] [PubMed] [Google Scholar]
  19. Lee S. S., Weiss R. S., Javier R. T. Binding of human virus oncoproteins to hDlg/SAP97, a mammalian homolog of the Drosophila discs large tumor suppressor protein. Proc Natl Acad Sci U S A. 1997 Jun 24;94(13):6670–6675. doi: 10.1073/pnas.94.13.6670. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Leslie N. R., Gray A., Pass I., Orchiston E. A., Downes C. P. Analysis of the cellular functions of PTEN using catalytic domain and C-terminal mutations: differential effects of C-terminal deletion on signalling pathways downstream of phosphoinositide 3-kinase. Biochem J. 2000 Mar 15;346(Pt 3):827–833. [PMC free article] [PubMed] [Google Scholar]
  21. Li D. M., Sun H. PTEN/MMAC1/TEP1 suppresses the tumorigenicity and induces G1 cell cycle arrest in human glioblastoma cells. Proc Natl Acad Sci U S A. 1998 Dec 22;95(26):15406–15411. doi: 10.1073/pnas.95.26.15406. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Li D. M., Sun H. TEP1, encoded by a candidate tumor suppressor locus, is a novel protein tyrosine phosphatase regulated by transforming growth factor beta. Cancer Res. 1997 Jun 1;57(11):2124–2129. [PubMed] [Google Scholar]
  23. Liliental J., Moon S. Y., Lesche R., Mamillapalli R., Li D., Zheng Y., Sun H., Wu H. Genetic deletion of the Pten tumor suppressor gene promotes cell motility by activation of Rac1 and Cdc42 GTPases. Curr Biol. 2000 Apr 6;10(7):401–404. doi: 10.1016/s0960-9822(00)00417-6. [DOI] [PubMed] [Google Scholar]
  24. Maehama T., Dixon J. E. PTEN: a tumour suppressor that functions as a phospholipid phosphatase. Trends Cell Biol. 1999 Apr;9(4):125–128. doi: 10.1016/s0962-8924(99)01519-6. [DOI] [PubMed] [Google Scholar]
  25. Maehama T., Dixon J. E. The tumor suppressor, PTEN/MMAC1, dephosphorylates the lipid second messenger, phosphatidylinositol 3,4,5-trisphosphate. J Biol Chem. 1998 May 29;273(22):13375–13378. doi: 10.1074/jbc.273.22.13375. [DOI] [PubMed] [Google Scholar]
  26. McCabe J. B., Berthiaume L. G. Functional roles for fatty acylated amino-terminal domains in subcellular localization. Mol Biol Cell. 1999 Nov;10(11):3771–3786. doi: 10.1091/mbc.10.11.3771. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Myers M. P., Pass I., Batty I. H., Van der Kaay J., Stolarov J. P., Hemmings B. A., Wigler M. H., Downes C. P., Tonks N. K. The lipid phosphatase activity of PTEN is critical for its tumor supressor function. Proc Natl Acad Sci U S A. 1998 Nov 10;95(23):13513–13518. doi: 10.1073/pnas.95.23.13513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Myers M. P., Stolarov J. P., Eng C., Li J., Wang S. I., Wigler M. H., Parsons R., Tonks N. K. P-TEN, the tumor suppressor from human chromosome 10q23, is a dual-specificity phosphatase. Proc Natl Acad Sci U S A. 1997 Aug 19;94(17):9052–9057. doi: 10.1073/pnas.94.17.9052. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Nakamura N., Ramaswamy S., Vazquez F., Signoretti S., Loda M., Sellers W. R. Forkhead transcription factors are critical effectors of cell death and cell cycle arrest downstream of PTEN. Mol Cell Biol. 2000 Dec;20(23):8969–8982. doi: 10.1128/mcb.20.23.8969-8982.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Ogg S., Ruvkun G. The C. elegans PTEN homolog, DAF-18, acts in the insulin receptor-like metabolic signaling pathway. Mol Cell. 1998 Dec;2(6):887–893. doi: 10.1016/s1097-2765(00)80303-2. [DOI] [PubMed] [Google Scholar]
  31. Ramaswamy S., Nakamura N., Vazquez F., Batt D. B., Perera S., Roberts T. M., Sellers W. R. Regulation of G1 progression by the PTEN tumor suppressor protein is linked to inhibition of the phosphatidylinositol 3-kinase/Akt pathway. Proc Natl Acad Sci U S A. 1999 Mar 2;96(5):2110–2115. doi: 10.1073/pnas.96.5.2110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Servant G., Weiner O. D., Herzmark P., Balla T., Sedat J. W., Bourne H. R. Polarization of chemoattractant receptor signaling during neutrophil chemotaxis. Science. 2000 Feb 11;287(5455):1037–1040. doi: 10.1126/science.287.5455.1037. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Shaw L. M., Rabinovitz I., Wang H. H., Toker A., Mercurio A. M. Activation of phosphoinositide 3-OH kinase by the alpha6beta4 integrin promotes carcinoma invasion. Cell. 1997 Dec 26;91(7):949–960. doi: 10.1016/s0092-8674(00)80486-9. [DOI] [PubMed] [Google Scholar]
  34. Shaw M., Cohen P. Role of protein kinase B and the MAP kinase cascade in mediating the EGF-dependent inhibition of glycogen synthase kinase 3 in Swiss 3T3 cells. FEBS Lett. 1999 Nov 12;461(1-2):120–124. doi: 10.1016/s0014-5793(99)01434-9. [DOI] [PubMed] [Google Scholar]
  35. Stambolic V., Suzuki A., de la Pompa J. L., Brothers G. M., Mirtsos C., Sasaki T., Ruland J., Penninger J. M., Siderovski D. P., Mak T. W. Negative regulation of PKB/Akt-dependent cell survival by the tumor suppressor PTEN. Cell. 1998 Oct 2;95(1):29–39. doi: 10.1016/s0092-8674(00)81780-8. [DOI] [PubMed] [Google Scholar]
  36. Sun H., Lesche R., Li D. M., Liliental J., Zhang H., Gao J., Gavrilova N., Mueller B., Liu X., Wu H. PTEN modulates cell cycle progression and cell survival by regulating phosphatidylinositol 3,4,5,-trisphosphate and Akt/protein kinase B signaling pathway. Proc Natl Acad Sci U S A. 1999 May 25;96(11):6199–6204. doi: 10.1073/pnas.96.11.6199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Tamura M., Gu J., Matsumoto K., Aota S., Parsons R., Yamada K. M. Inhibition of cell migration, spreading, and focal adhesions by tumor suppressor PTEN. Science. 1998 Jun 5;280(5369):1614–1617. doi: 10.1126/science.280.5369.1614. [DOI] [PubMed] [Google Scholar]
  38. Tamura M., Gu J., Takino T., Yamada K. M. Tumor suppressor PTEN inhibition of cell invasion, migration, and growth: differential involvement of focal adhesion kinase and p130Cas. Cancer Res. 1999 Jan 15;59(2):442–449. [PubMed] [Google Scholar]
  39. Taylor V., Wong M., Brandts C., Reilly L., Dean N. M., Cowsert L. M., Moodie S., Stokoe D. 5' phospholipid phosphatase SHIP-2 causes protein kinase B inactivation and cell cycle arrest in glioblastoma cells. Mol Cell Biol. 2000 Sep;20(18):6860–6871. doi: 10.1128/mcb.20.18.6860-6871.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Torres J., Pulido R. The tumor suppressor PTEN is phosphorylated by the protein kinase CK2 at its C terminus. Implications for PTEN stability to proteasome-mediated degradation. J Biol Chem. 2001 Jan 12;276(2):993–998. doi: 10.1074/jbc.M009134200. [DOI] [PubMed] [Google Scholar]
  41. Vanhaesebroeck B., Alessi D. R. The PI3K-PDK1 connection: more than just a road to PKB. Biochem J. 2000 Mar 15;346(Pt 3):561–576. [PMC free article] [PubMed] [Google Scholar]
  42. Vazquez F., Ramaswamy S., Nakamura N., Sellers W. R. Phosphorylation of the PTEN tail regulates protein stability and function. Mol Cell Biol. 2000 Jul;20(14):5010–5018. doi: 10.1128/mcb.20.14.5010-5018.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Watton S. J., Downward J. Akt/PKB localisation and 3' phosphoinositide generation at sites of epithelial cell-matrix and cell-cell interaction. Curr Biol. 1999 Apr 22;9(8):433–436. doi: 10.1016/s0960-9822(99)80192-4. [DOI] [PubMed] [Google Scholar]
  44. Welch H., Eguinoa A., Stephens L. R., Hawkins P. T. Protein kinase B and rac are activated in parallel within a phosphatidylinositide 3OH-kinase-controlled signaling pathway. J Biol Chem. 1998 May 1;273(18):11248–11256. doi: 10.1074/jbc.273.18.11248. [DOI] [PubMed] [Google Scholar]
  45. Wennström S., Hawkins P., Cooke F., Hara K., Yonezawa K., Kasuga M., Jackson T., Claesson-Welsh L., Stephens L. Activation of phosphoinositide 3-kinase is required for PDGF-stimulated membrane ruffling. Curr Biol. 1994 May 1;4(5):385–393. doi: 10.1016/s0960-9822(00)00087-7. [DOI] [PubMed] [Google Scholar]
  46. Williams M. R., Arthur J. S., Balendran A., van der Kaay J., Poli V., Cohen P., Alessi D. R. The role of 3-phosphoinositide-dependent protein kinase 1 in activating AGC kinases defined in embryonic stem cells. Curr Biol. 2000 Apr 20;10(8):439–448. doi: 10.1016/s0960-9822(00)00441-3. [DOI] [PubMed] [Google Scholar]
  47. Wu X., Hepner K., Castelino-Prabhu S., Do D., Kaye M. B., Yuan X. J., Wood J., Ross C., Sawyers C. L., Whang Y. E. Evidence for regulation of the PTEN tumor suppressor by a membrane-localized multi-PDZ domain containing scaffold protein MAGI-2. Proc Natl Acad Sci U S A. 2000 Apr 11;97(8):4233–4238. doi: 10.1073/pnas.97.8.4233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wu Y., Dowbenko D., Spencer S., Laura R., Lee J., Gu Q., Lasky L. A. Interaction of the tumor suppressor PTEN/MMAC with a PDZ domain of MAGI3, a novel membrane-associated guanylate kinase. J Biol Chem. 2000 Jul 14;275(28):21477–21485. doi: 10.1074/jbc.M909741199. [DOI] [PubMed] [Google Scholar]
  49. Zhou X. P., Li Y. J., Hoang-Xuan K., Laurent-Puig P., Mokhtari K., Longy M., Sanson M., Delattre J. Y., Thomas G., Hamelin R. Mutational analysis of the PTEN gene in gliomas: molecular and pathological correlations. Int J Cancer. 1999 Apr 20;84(2):150–154. doi: 10.1002/(sici)1097-0215(19990420)84:2<150::aid-ijc10>3.0.co;2-#. [DOI] [PubMed] [Google Scholar]
  50. van Weering D. H., de Rooij J., Marte B., Downward J., Bos J. L., Burgering B. M. Protein kinase B activation and lamellipodium formation are independent phosphoinositide 3-kinase-mediated events differentially regulated by endogenous Ras. Mol Cell Biol. 1998 Apr;18(4):1802–1811. doi: 10.1128/mcb.18.4.1802. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. van der Kaay J., Batty I. H., Cross D. A., Watt P. W., Downes C. P. A novel, rapid, and highly sensitive mass assay for phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) and its application to measure insulin-stimulated PtdIns(3,4,5)P3 production in rat skeletal muscle in vivo. J Biol Chem. 1997 Feb 28;272(9):5477–5481. doi: 10.1074/jbc.272.9.5477. [DOI] [PubMed] [Google Scholar]

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