Skip to main content

Pharmacological Inhibition of Endocytic Pathways: Is It Specific Enough to Be Useful?

  • Protocol
Exocytosis and Endocytosis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 440))

Summary

Eukaryotic cells constantly form and internalize plasma membrane vesicles in a process known as endocytosis. Endocytosis serves a variety of housekeeping and specialized cellular functions, and it can be mediated by distinct molecular pathways. Among them, internalization via clathrin-coated pits, lipid raft/caveolae-mediated endocytosis and macropinocytosis/phagocytosis are the most extensively characterized. The major endocytic pathways are usually distinguished on the basis of their differential sensitivity to pharmacological/chemical inhibitors, although the possibility of nonspecific effects of such inhibitors is frequently overlooked. This review provides a critical evaluation of the selectivity of the most widely used pharmacological inhibitors of clathrin-mediated, lipid raft/caveolae-mediated endocytosis and macropinocytosis/phagocytosis. The mechanisms of actions of these agents are described with special emphasis on their reported side effects on the alternative internalization modes and the actin cytoskeleton. The most and the least-selective inhibitors of each major endocytic pathway are highlighted.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. 1. Conner, S.D., and Schmid, S.L. (2003) Regulated portals of entry into the cell. Nature 422, 37–44.

    Article  CAS  PubMed  Google Scholar 

  2. 2. Marsh, M., and McMahon, H.T. (1999) The structural era of endocytosis. Science 285, 215–220.

    Article  CAS  PubMed  Google Scholar 

  3. 3. Parton, R.G., and Richards, A.A. (2003) Lipid rafts and caveolae as portals for endocytosis: new insights and common mechanisms. Traffic 4, 724–738.

    Article  CAS  PubMed  Google Scholar 

  4. 4. Amyere, M., Mettlen, M., Van Der Smissen, P., et al. (2002) Origin, originality, functions, subversions and molecular signalling of macropinocytosis. Int. J. Med. Microbiol. 291, 487–494.

    Article  CAS  PubMed  Google Scholar 

  5. 5. Niedergang, F., and Chavrier, P. (2004) Signaling and membrane dynamics during phagocytosis: many roads lead to the phagos(R)ome. Curr. Opin. Cell Biol. 16, 422–428.

    Article  CAS  PubMed  Google Scholar 

  6. 6. Swanson, J.A., and Watts, C. (1995) Macropinocytosis. Trends Cell Biol. 5, 424–428.

    Article  CAS  PubMed  Google Scholar 

  7. 7. Davies, P.J., Davies, D.R., Levitzki, A., et al. (1980) Transglutaminase is essential in receptor-mediated endocytosis of alpha 2-macroglobulin and polypeptide hormones. Nature 283, 162–167.

    Article  CAS  PubMed  Google Scholar 

  8. 8. Larkin, J.M., Brown, M.S., Goldstein, J.L., and Anderson, R.G. (1983) Depletion of intracellular potassium arrests coated pit formation and receptor-mediated endocytosis in fibroblasts. Cell 33, 273–285.

    Article  CAS  PubMed  Google Scholar 

  9. 9. Daukas, G., and Zigmond, S.H. (1985) Inhibition of receptor-mediated but not fluid-phase endocytosis in polymorphonuclear leukocytes. J. Cell Biol. 101, 1673–1679.

    Article  CAS  PubMed  Google Scholar 

  10. 10. Hansen, S.H., Sandvig, K., and van Deurs, B. (1993) Clathrin and HA2 adaptors: effects of potassium depletion, hypertonic medium, and cytosol acidification. J. Cell Biol. 121, 61–72.

    Article  CAS  PubMed  Google Scholar 

  11. 11. Inal, J., Miot, S., and Schifferli, J.A. (2005) The complement inhibitor, CRIT, undergoes clathrin-dependent endocytosis. Exp. Cell Res. 310, 54–65.

    Article  CAS  PubMed  Google Scholar 

  12. 12. Tulapurkar, M.E., Schafer, R., Hanck, T., et al. (2005) Endocytosis mechanism of P2Y2 nucleotide receptor tagged with green fluorescent protein: clathrin and actin cytoskeleton dependence. Cell. Mol. Life Sci. 62, 1388–1399.

    Article  CAS  PubMed  Google Scholar 

  13. 13. Yao, D., Ehrlich, M., Henis, Y.I., and Leof, E.B. (2002) Transforming growth factor-β receptors interact with AP2 by direct binding to β2 subunit. Mol. Biol. Cell 13, 4001–4012.

    Article  CAS  PubMed  Google Scholar 

  14. 14. Carpentier, J.L., Sawano, F., Geiger, D., et al. (1989) Potassium depletion and hypertonic medium reduce “non-coated” and clathrin-coated pit formation, as well as endocytosis through these two gates. J. Cell Physiol. 138, 519–526.

    Article  CAS  PubMed  Google Scholar 

  15. 15. Bradley, J.R., Johnson, D.R., and Pober, J.S. (1993) Four different classes of inhibitors of receptor-mediated endocytosis decrease tumor necrosis factor-induced gene expression in human endothelial cells. J. Immunol. 150, 5544–5555.

    CAS  PubMed  Google Scholar 

  16. 16. Synnes, M., Prydz, K., Lovdal, T., Brech, A., and Berg, T. (1999) Fluid phase endocytosis and galactosyl receptor-mediated endocytosis employ different early endosomes. Biochim. Biophys. Acta 1421, 317–328.

    Article  CAS  PubMed  Google Scholar 

  17. 17. Page, E., Winterfield, J., Goings, G., Bastawrous, A., and Upshaw-Earley, J. (1998) Water channel proteins in rat cardiac myocyte caveolae: osmolarity-dependent reversible internalization. Am. J. Physiol. 274, H1988–H2000.

    CAS  PubMed  Google Scholar 

  18. 18. Malek, A.M., Xu, C., Kim, E.S., and Alper, S.L. (2006) Hypertonicity triggers RhoA-dependent assembly of myosin-containing striated polygonal actin networks in endothelial cells. Am. J. Physiol. Cell Physiol. 292, C1645–C1659.

    Article  PubMed  CAS  Google Scholar 

  19. 19. Bustamante, M., Roger, F., Bochaton-Piallat, M.L., et al. (2003) Regulatory volume increase is associated with p38 kinase-dependent actin cytoskeleton remodeling in rat kidney MTAL. Am. J. Physiol. Renal Physiol. 285, F336–F347.

    CAS  PubMed  Google Scholar 

  20. 20. Liu, J., Kesiry, R., Periyasamy, S.M., et al. (2004) Ouabain induces endocytosis of plasmalemmal Na/K-ATPase in LLC-PK1 cells by a clathrin-dependent mechanism. Kidney Int. 66, 227–241.

    Article  CAS  PubMed  Google Scholar 

  21. 21. Idkowiak-Baldys, J., Becker, K.P., Kitatani, K., and Hannun, Y.A. (2006) Dynamic sequestration of the recycling compartment by classical protein kinase C. J. Biol. Chem. 281, 22321–22331.

    Article  CAS  PubMed  Google Scholar 

  22. 22. Cupers, P., Veithen, A., Kiss, A., Baudhuin, P., and Courtoy, P.J. (1994) Clathrin polymerization is not required for bulk-phase endocytosis in rat fetal fibroblasts. J. Cell Biol. 127, 725–735.

    Article  CAS  PubMed  Google Scholar 

  23. 23. Yumoto, R., Nishikawa, H., Okamoto, M., et al. (2006) Clathrin-mediated endocytosis of FITC-albumin in alveolar type II epithelial cell line RLE-6TN. Am. J. Physiol. Lung Cell Mol. Physiol. 290, L946–L955.

    Article  CAS  PubMed  Google Scholar 

  24. 24. Altankov, G., and Grinnell, F. (1993) Depletion of intracellular potassium disrupts coated pits and reversibly inhibits cell polarization during fibroblast spreading. J. Cell Biol. 120, 1449–1459.

    Article  CAS  PubMed  Google Scholar 

  25. 25. Rajasekaran, S.A., Palmer, L.G., Moon, S.Y., et al. (2001) Na,K-ATPase activity is required for formation of tight junctions, desmosomes, and induction of polarity in epithelial cells. Mol. Biol. Cell 12, 3717–3732.

    CAS  PubMed  Google Scholar 

  26. 26. Cosson, P., de Curtis, I., Pouyssegur, J., Griffiths, G., and Davoust, J. (1989) Low cytoplasmic pH inhibits endocytosis and transport from the trans-Golgi network to the cell surface. J. Cell Biol. 108, 377–387.

    Article  CAS  PubMed  Google Scholar 

  27. 27. Ivanov, A.I., Nusrat, A., and Parkos, C.A. (2004) Endocytosis of epithelial apical junctional proteins by a clathrin-mediated pathway into a unique storage compartment. Mol. Biol. Cell 15, 176–188.

    Article  CAS  PubMed  Google Scholar 

  28. 28. Sandvig, K., Olsnes, S., Petersen, O.W., and van Deurs, B. (1987) Acidification of the cytosol inhibits endocytosis from coated pits. J. Cell Biol. 105, 679–689.

    Article  CAS  PubMed  Google Scholar 

  29. 29. Eker, P., Holm, P.K., van Deurs, B., and Sandvig, K. (1994) Selective regulation of apical endocytosis in polarized Madin–Darby canine kidney cells by mastoparan and cAMP. J. Biol. Chem. 269, 18607–18615.

    CAS  PubMed  Google Scholar 

  30. 30. Suzuki, K., and Namiki, H. (2007) Cytoplasmic pH-dependent spreading of polymorphonuclear leukocytes: regulation by pH of PKC subcellular distribution and F-actin assembly. Cell Biol. Int. 31, 279–288.

    Article  CAS  PubMed  Google Scholar 

  31. 31. Huotari, V., Vaaraniemi, J., Lehto, V.P., and Eskelinen, S. (1996) Regulation of the disassembly/assembly of the membrane skeleton in Madin–Darby canine kidney cells. J. Cell Physiol. 167, 121–130.

    Article  CAS  PubMed  Google Scholar 

  32. 32. Wang, L.H., Rothberg, K.G., and Anderson, R.G. (1993) Mis-assembly of clathrin lattices on endosomes reveals a regulatory switch for coated pit formation. J. Cell Biol. 123, 1107–1117.

    Article  CAS  PubMed  Google Scholar 

  33. 33. Elferink, J.G. (1979) Chlorpromazine inhibits phagocytosis and exocytosis in rabbit polymorphonuclear leukocytes. Biochem. Pharmacol. 28, 965–968.

    Article  CAS  PubMed  Google Scholar 

  34. 34. Watanabe, S., Hirose, M., Miyazaki, A., et al. (1988) Calmodulin antagonists inhibit the phagocytic activity of cultured Kupffer cells. Lab. Invest. 59, 214–218.

    CAS  PubMed  Google Scholar 

  35. 35. Ogiso, T., Iwaki, M., and Mori, K. (1981) Fluidity of human erythrocyte membrane and effect of chlorpromazine on fluidity and phase separation of membrane. Biochim. Biophys. Acta 649, 325–335.

    Article  CAS  PubMed  Google Scholar 

  36. 36. Giocondi, M.C., Mamdouh, Z., and Le Grimellec, C. (1995) Benzyl alcohol differently affects fluid phase endocytosis and exocytosis in renal epithelial cells. Biochim. Biophys. Acta 1234, 197–202.

    Article  PubMed  Google Scholar 

  37. 37. Walenga, R.W., Opas, E.E., and Feinstein, M.B. (1981) Differential effects of calmodulin antagonists on phospholipases A2 and C in thrombin-stimulated platelets. J. Biol. Chem. 256, 12523–12528.

    CAS  PubMed  Google Scholar 

  38. 38. Wells, A., Ware, M.F., Allen, F.D., and Lauffenburger, D.A. (1999) Shaping up for shipping out: PLCγ signaling of morphology changes in EGF-stimulated fibroblast migration. Cell Motil. Cytoskeleton 44, 227–233.

    Article  CAS  PubMed  Google Scholar 

  39. 39. Amyere, M., Payrastre, B., Krause, U., et al. (2000) Constitutive macropinocytosis in oncogene-transformed fibroblasts depends on sequential permanent activation of phosphoinositide 3-kinase and phospholipase C. Mol. Biol. Cell 11, 3453–3467.

    CAS  PubMed  Google Scholar 

  40. 40. Veithen, A., Cupers, P., Baudhuin, P., and Courtoy, P.J. (1996) v-Src induces constitutive macropinocytosis in rat fibroblasts. J. Cell Sci. 109, 2005–2012.

    CAS  PubMed  Google Scholar 

  41. 41. Panicker, A.K., Buhusi, M., Erickson, A., and Maness, P.F. (2006) Endocytosis of β1 integrins is an early event in migration promoted by the cell adhesion molecule L1. Exp. Cell Res. 312, 299–307.

    CAS  PubMed  Google Scholar 

  42. 42. Wang, J., and Liu, X.J. (2003) A G protein-coupled receptor kinase induces Xenopus oocyte maturation. J. Biol. Chem. 278, 15809–15814.

    Article  CAS  PubMed  Google Scholar 

  43. 43. Nandi, P.K., Van Jaarsveld, P.P., Lippoldt, R.E., and Edelhoch, H. (1981) Effect of basic compounds on the polymerization of clathrin. Biochemistry 20, 6706–6710.

    Article  CAS  PubMed  Google Scholar 

  44. 44. Leu, R.W., Herriott, M.J., Moore, P.E., et al. (1982) Enhanced transglutaminase activity associated with macrophage activation. Possible role in Fc-mediated phagocytosis. Exp. Cell Res. 141, 191–199.

    Article  CAS  PubMed  Google Scholar 

  45. 45. Thompson, K., Rogers, M.J., Coxon, F.P., and Crockett, J.C. (2006) Cytosolic entry of bisphosphonate drugs requires acidification of vesicles after fluid-phase endocytosis. Mol. Pharmacol. 69, 1624–1632.

    Article  CAS  PubMed  Google Scholar 

  46. 46. Schlegel, R., Dickson, R.B., Willingham, M.C., and Pastan, I.H. (1982) Amantadine and dansylcadaverine inhibit vesicular stomatitis virus uptake and receptor-mediated endocytosis of alpha 2-macroglobulin. Proc. Natl. Acad. Sci. U. S. A. 79, 2291–2295.

    Article  CAS  PubMed  Google Scholar 

  47. 47. Mishra, S., and Murphy, L.J. (2004) Tissue transglutaminase has intrinsic kinase activity: identification of transglutaminase 2 as an insulin-like growth factor-binding protein-3 kinase. J. Biol. Chem. 279, 23863–23868.

    Article  CAS  PubMed  Google Scholar 

  48. 48. Singh, U.S., Pan, J., Kao, Y.L., et al. (2003) Tissue transglutaminase mediates activation of RhoA and MAP kinase pathways during retinoic acid-induced neuronal differentiation of SH-SY5Y cells. J. Biol. Chem. 278, 391–399.

    Article  CAS  PubMed  Google Scholar 

  49. 49. Kang, S.J., Shin, K.S., Song, W.K., et al. (1995) Involvement of transglutaminase in myofibril assembly of chick embryonic myoblasts in culture. J. Cell Biol. 130, 1127–1136.

    Article  CAS  PubMed  Google Scholar 

  50. 50. Gibson, A.E., Noel, R.J., Herlihy, J.T., and Ward, W.F. (1989) Phenylarsine oxide inhibition of endocytosis: effects on asialofetuin internalization. Am. J. Physiol. 257, C182–C184.

    CAS  PubMed  Google Scholar 

  51. 51. Sturrock, A., Alexander, J., Lamb, J., et al. (1990) Characterization of a transferrin-independent uptake system for iron in HeLa cells. J. Biol. Chem. 265, 3139–3145.

    CAS  PubMed  Google Scholar 

  52. 52. Frost, S.C., Lane, M.D., and Gibbs, E.M. (1989) Effect of phenylarsine oxide on fluid phase endocytosis: further evidence for activation of the glucose transporter. J. Cell Physiol. 141, 467–474.

    Article  CAS  PubMed  Google Scholar 

  53. 53. Massol, P., Montcourrier, P., Guillemot, J.C., and Chavrier, P. (1998) Fc receptor-mediated phagocytosis requires Cdc42 and Rac1. EMBO J. 17, 6219–6229.

    Article  CAS  PubMed  Google Scholar 

  54. 54. Frost, S.C., and Lane, M.D. (1985) Evidence for the involvement of vicinal sulfhydryl groups in insulin-activated hexose transport by 3T3-L1 adipocytes. J. Biol. Chem. 260, 2646–2652.

    CAS  PubMed  Google Scholar 

  55. 55. Retta, S.F., Barry, S.T., Critchley, D.R., et al. (1996) Focal adhesion and stress fiber formation is regulated by tyrosine phosphatase activity. Exp. Cell Res. 229, 307–317.

    Article  CAS  PubMed  Google Scholar 

  56. 56. Gerhard, R., John, H., Aktories, K., and Just, I. (2003) Thiol-modifying phenylarsine oxide inhibits guanine nucleotide binding of Rho but not of Rac GTPases. Mol. Pharmacol. 63, 1349–1355.

    Article  CAS  PubMed  Google Scholar 

  57. 57. Smart, E.J., and Anderson, R.G. (2002) Alterations in membrane cholesterol that affect structure and function of caveolae. Methods Enzymol. 353, 131–139.

    Article  CAS  PubMed  Google Scholar 

  58. 58. Liao, J.K., and Laufs, U. (2005) Pleiotropic effects of statins. Annu. Rev. Pharmacol. Toxicol. 45, 89–118.

    Article  CAS  PubMed  Google Scholar 

  59. 59. Tobert, J.A. (2003) Lovastatin and beyond: the history of the HMG-CoA reductase inhibitors. Nat. Rev. Drug Discov. 2, 517–526.

    Article  CAS  PubMed  Google Scholar 

  60. 60. Sidaway, J.E., Davidson, R.G., McTaggart, F., et al. (2004) Inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase reduce receptor-mediated endocytosis in opossum kidney cells. J. Am. Soc. Nephrol. 15, 2258–2265.

    Article  CAS  PubMed  Google Scholar 

  61. 61. Chan, P.C., Lafreniere, R., and Parsons, H.G. (1997) Lovastatin increases surface low density lipoprotein receptor expression by retarding the receptor internalization rate in proliferating lymphocytes. Biochem. Biophys. Res. Commun. 235, 117–122.

    Article  CAS  PubMed  Google Scholar 

  62. 62. Loike, J.D., Shabtai, D.Y., Neuhut, R., et al. (2004) Statin inhibition of Fc receptor-mediated phagocytosis by macrophages is modulated by cell activation and cholesterol. Arterioscler. Thromb. Vasc. Biol. 24, 2051–2056.

    Article  CAS  PubMed  Google Scholar 

  63. 63. Desnoyers, L., Anant, J.S., and Seabra, M.C. (1996) Geranylgeranylation of Rab proteins. Biochem. Soc. Trans. 24, 699–703.

    CAS  PubMed  Google Scholar 

  64. 64. Cordle, A., Koenigsknecht-Talboo, J., Wilkinson, B., et al. (2005) Mechanisms of statin-mediated inhibition of small G-protein function. J. Biol. Chem. 280, 34202–34209.

    Article  CAS  PubMed  Google Scholar 

  65. 65. Irie, T., Fukunaga, K., and Pitha, J. (1992) Hydroxypropylcyclodextrins in parenteral use. I: Lipid dissolution and effects on lipid transfers in vitro. J. Pharm. Sci. 81, 521–523.

    Article  CAS  PubMed  Google Scholar 

  66. 66. Kilsdonk, E.P., Yancey, P.G., Stoudt, G.W., et al. (1995) Cellular cholesterol efflux mediated by cyclodextrins. J. Biol. Chem. 270, 17250–17256.

    Article  CAS  PubMed  Google Scholar 

  67. 67. Westermann, M., Steiniger, F., and Richter, W. (2005) Belt-like localization of caveolin in deep caveolae and its re-distribution after cholesterol depletion. Histochem. Cell Biol. 123, 613–620.

    Article  CAS  PubMed  Google Scholar 

  68. 68. Lu, L., Khan, S., Lencer, W., and Walker, W.A. (2005) Endocytosis of cholera toxin by human enterocytes is developmentally regulated. Am. J. Physiol. Gastrointest. Liver Physiol. 289, G332–G341.

    Article  CAS  PubMed  Google Scholar 

  69. 69. Shigematsu, S., Watson, R.T., Khan, A.H., and Pessin, J.E. (2003) The adipocyte plasma membrane caveolin functional/structural organization is necessary for the efficient endocytosis of GLUT4. J. Biol. Chem. 278, 10683–10690.

    Article  CAS  PubMed  Google Scholar 

  70. 70. Maniatis, N.A., Brovkovych, V., Allen, S.E., et al. (2006) Novel mechanism of endothelial nitric oxide synthase activation mediated by caveolae internalization in endothelial cells. Circ. Res. 99, 870–877.

    Article  CAS  PubMed  Google Scholar 

  71. 71. Rodal, S.K., Skretting, G., Garred, O., et al. (1999) Extraction of cholesterol with methyl-beta-cyclodextrin perturbs formation of clathrin-coated endocytic vesicles. Mol. Biol. Cell 10, 961–974.

    CAS  PubMed  Google Scholar 

  72. 72. Liu, N.Q., Lossinsky, A.S., Popik, et al. (2002) Human immunodeficiency virus type 1 enters brain microvascular endothelia by macropinocytosis dependent on lipid rafts and the mitogen-activated protein kinase signaling pathway. J. Virol. 76, 6689–6700.

    Article  CAS  PubMed  Google Scholar 

  73. 73. Lu, H., Sun, T.X., Bouley, R., et al. (2004) Inhibition of endocytosis causes phosphorylation (S256)-independent plasma membrane accumulation of AQP2. Am. J. Physiol. Renal Physiol. 286, F233–F243.

    Article  CAS  PubMed  Google Scholar 

  74. 74. Kanzaki, M., and Pessin, J.E. (2002) Caveolin-associated filamentous actin (Cav-actin) defines a novel F-actin structure in adipocytes. J. Biol. Chem. 277, 25867–25869.

    Article  CAS  PubMed  Google Scholar 

  75. 75. Pike, L.J., and Miller, J.M. (1998) Cholesterol depletion delocalizes phosphatidylinositol bisphosphate and inhibits hormone-stimulated phosphatidylinositol turnover. J. Biol. Chem. 273, 22298–22304.

    Article  CAS  PubMed  Google Scholar 

  76. 76. Kranenburg, O., Verlaan, I., and Moolenaar, W.H. (2001) Regulating c-Ras function. cholesterol depletion affects caveolin association, GTP loading, and signaling. Curr. Biol. 11, 1880–1884.

    Article  CAS  PubMed  Google Scholar 

  77. 77. Kitajima, Y., Sekiya, T., and Nozawa, Y. (1976) Freeze-fracture ultrastructural alterations induced by filipin, pimaricin, nystatin and amphotericin B in the plasmia membranes of Epidermophyton, Saccharomyces and red complex-induced membrane lesions. Biochim. Biophys. Acta 455, 452–465.

    Article  CAS  PubMed  Google Scholar 

  78. 78. Ros-Baro, A., Lopez-Iglesias, C., Peiro, S., et al. (2001) Lipid rafts are required for GLUT4 internalization in adipose cells. Proc. Natl. Acad. Sci. U. S. A. 98, 12050–12055.

    Article  CAS  PubMed  Google Scholar 

  79. 79. Rothberg, K.G., Ying, Y.S., Kamen, B.A., and Anderson, R.G. (1990) Cholesterol controls the clustering of the glycophospholipid-anchored membrane receptor for 5-methyltetrahydrofolate. J. Cell Biol. 111, 2931–2938.

    Article  CAS  PubMed  Google Scholar 

  80. 80. Orlandi, P.A., and Fishman, P.H. (1998) Filipin-dependent inhibition of cholera toxin: evidence for toxin internalization and activation through caveolae-like domains. J. Cell Biol. 141, 905–915.

    Article  CAS  PubMed  Google Scholar 

  81. 81. Singh, R.D., Puri, V., Valiyaveettil, J.T., et al. (2003) Selective caveolin-1-dependent endocytosis of glycosphingolipids. Mol. Biol. Cell 14, 3254–3265.

    Article  CAS  PubMed  Google Scholar 

  82. 82. Monis, G.F., Schultz, C., Ren, R., et al. (2006) Role of endocytic inhibitory drugs on internalization of amyloidogenic light chains by cardiac fibroblasts. Am. J. Pathol. 169, 1939–1952.

    Article  CAS  PubMed  Google Scholar 

  83. 83. Milhaud, J. (1992) Permeabilizing action of filipin III on model membranes through a filipin-phospholipid binding. Biochim. Biophys. Acta 1105, 307–318.

    Article  CAS  PubMed  Google Scholar 

  84. 84. Harder, T., Kellner, R., Parton, R.G., and Gruenberg, J. (1997) Specific release of membrane-bound annexin II and cortical cytoskeletal elements by sequestration of membrane cholesterol. Mol. Biol. Cell 8, 533–545.

    CAS  PubMed  Google Scholar 

  85. 85. MacLachlan, J., Wotherspoon, A.T., Ansell, R.O., and Brooks, C.J. (2000) Cholesterol oxidase: sources, physical properties and analytical applications. J. Steroid Biochem. Mol. Biol. 72, 169–195.

    Article  CAS  PubMed  Google Scholar 

  86. 86. Smart, E.J., Ying, Y.S., Conrad, P.A., and Anderson, R.G. (1994) Caveolin moves from caveolae to the Golgi apparatus in response to cholesterol oxidation. J. Cell Biol. 127, 1185–1197.

    Article  CAS  PubMed  Google Scholar 

  87. 87. Thyberg, J. (2003) Cholesterol oxidase and the hydroxymethylglutaryl coenzyme A reductase inhibitor mevinolin perturb endocytic trafficking in cultured vascular smooth muscle cells. J. Submicrosc. Cytol. Pathol. 35, 457–468.

    CAS  PubMed  Google Scholar 

  88. 88. Brasaemle, D.L., and Attie, A.D. (1990) Rapid intracellular transport of LDL-derived cholesterol to the plasma membrane in cultured fibroblasts. J. Lipid Res. 31, 103–112.

    CAS  PubMed  Google Scholar 

  89. 89. Lange, Y. (1992) Tracking cell cholesterol with cholesterol oxidase. J. Lipid Res. 33, 315–321.

    CAS  PubMed  Google Scholar 

  90. 90. Okamoto, Y., Ninomiya, H., Miwa, S., and Masaki, T. (2000) Cholesterol oxidation switches the internalization pathway of endothelin receptor type A from caveolae to clathrin-coated pits in Chinese hamster ovary cells. J. Biol. Chem. 275, 6439–6446.

    Article  CAS  PubMed  Google Scholar 

  91. 91. Piehl, M., Lehmann, C., Gumpert, A., et al. (2007) Internalization of large double-membrane intercellular vesicles by a clathrin-dependent endocytic process. Mol. Biol. Cell 18, 337–347.

    Article  CAS  PubMed  Google Scholar 

  92. 92. Veiga, E., and Cossart, P. (2006) The role of clathrin-dependent endocytosis in bacterial internalization. Trends Cell Biol. 16, 499–504.

    Article  CAS  PubMed  Google Scholar 

  93. 93. West, M.A., Bretscher, M.S., and Watts, C. 1989. Distinct endocytotic pathways in epidermal growth factor-stimulated human carcinoma A431 cells. J. Cell Biol. 109, 2731–2739.

    Article  CAS  PubMed  Google Scholar 

  94. 94. Marechal, V., Prevost, M.C., Petit, C., et al. (2001) Human immunodeficiency virus type 1 entry into macrophages mediated by macropinocytosis. J. Virol. 75, 11166–11177.

    Article  CAS  PubMed  Google Scholar 

  95. 95. Nakase, I., Niwa, M., Takeuchi, T., et al. (2004) Cellular uptake of arginine-rich peptides: roles for macropinocytosis and actin rearrangement. Mol. Ther. 10, 1011–1022.

    Article  CAS  PubMed  Google Scholar 

  96. 96. von Delwig, A., Bailey, E., Gibbs, D.M., and Robinson, J.H. (2002) The route of bacterial uptake by macrophages influences the repertoire of epitopes presented to CD4 T cells. Eur. J. Immunol. 32, 3714–3719.

    Article  Google Scholar 

  97. 97. Fretz, M., Jin, J., Conibere, R., et al. (2006) Effects of Na+/H+ exchanger inhibitors on subcellular localisation of endocytic organelles and intracellular dynamics of protein transduction domains HIV-TAT peptide and octaarginine. J. Control Release 116, 247–254.

    Article  CAS  PubMed  Google Scholar 

  98. 98. Meier, O., Boucke, K., Hammer, S.V., et al. (2002) Adenovirus triggers macropinocytosis and endosomal leakage together with its clathrin-mediated uptake. J. Cell Biol. 158, 1119–1131.

    Article  CAS  PubMed  Google Scholar 

  99. 99. Wadia, J.S., Stan, R.V., and Dowdy, S.F. (2004) Transducible TAT-HA fusogenic peptide enhances escape of TAT-fusion proteins after lipid raft macropinocytosis. Nat. Med. 10, 310–315.

    Article  CAS  PubMed  Google Scholar 

  100. 100. Lagana, A., Vadnais, J., Le, P.U., et al. (2000) Regulation of the formation of tumor cell pseudopodia by the Na+/H+ exchanger NHE1. J. Cell Sci. 113, 3649–3662.

    CAS  PubMed  Google Scholar 

  101. 101. Peterson, J.R., and Mitchison, T.J. (2002) Small molecules, big impact: a history of chemical inhibitors and the cytoskeleton. Chem. Biol. 9, 1275–1285.

    Article  CAS  PubMed  Google Scholar 

  102. 102. Dharmawardhane, S., Schurmann, A., Sells, M.A., et al. 2000. Regulation of macropinocytosis by p21-activated kinase-1. Mol. Biol. Cell 11, 3341–3352.

    CAS  PubMed  Google Scholar 

  103. 103. Mettlen, M., Platek, A., Van Der Smissen, P., et al. (2006) Src triggers circular ruffling and macropinocytosis at the apical surface of polarized MDCK cells. Traffic 7:589–603.

    Article  CAS  Google Scholar 

  104. 104. Montaner, L.J., da Silva, R.P., Sun, J., et al. (1999) Type 1 and type 2 cytokine regulation of macrophage endocytosis: differential activation by IL-4/IL-13 as opposed to IFN-γ or IL-10. J. Immunol. 162, 4606–4613.

    CAS  PubMed  Google Scholar 

  105. 105. Kaksonen, M., Toret, C.P., and Drubin, D.G. (2006) Harnessing actin dynamics for clathrin-mediated endocytosis. Nat. Rev. Mol. Cell Biol. 7, 404–414.

    Article  CAS  PubMed  Google Scholar 

  106. 106. Merrifield, C.J. (2004) Seeing is believing: imaging actin dynamics at single sites of endocytosis. Trends Cell Biol. 14, 352–358.

    Article  CAS  PubMed  Google Scholar 

  107. 107. Takenawa, T., and Itoh, T. (2001) Phosphoinositides, key molecules for regulation of actin cytoskeletal organization and membrane traffic from the plasma membrane. Biochim. Biophys. Acta 1533, 190–206.

    CAS  PubMed  Google Scholar 

  108. 108. Araki, N., Johnson, M.T., and Swanson, J.A. (1996) A role for phosphoinositide 3-kinase in the completion of macropinocytosis and phagocytosis by macrophages. J. Cell Biol. 135, 1249–1260.

    Article  CAS  PubMed  Google Scholar 

  109. 109. Roth, M.G. (2004) Phosphoinositides in constitutive membrane traffic. Physiol. Rev. 84, 699–730.

    Article  CAS  PubMed  Google Scholar 

  110. 110. Jess, T.J., Belham, C.M., Thomson, F.J., (1996) Phosphatidylinositol 3′-kinase, but not p70 ribosomal S6 kinase, is involved in membrane protein recycling: wortmannin inhibits glucose transport and downregulates cell-surface transferrin receptor numbers independently of any effect on fluid-phase endocytosis in fibroblasts. Cell Signal. 8, 297–304.

    Article  CAS  PubMed  Google Scholar 

  111. 111. Hill, T., Odell, L.R., Edwards, J.K., et al. (2005) Small molecule inhibitors of dynamin I GTPase activity: development of dimeric tyrphostins. J. Med. Chem. 48, 7781–7788.

    Article  CAS  PubMed  Google Scholar 

  112. 112. Macia, E., Ehrlich, M., Massol, R., et al. (2006) Dynasore, a cell-permeable inhibitor of dynamin. Dev. Cell 10, 839–850.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

I thank Moshe Bachar for his excellent editorial assistance and Drs. Alex Mongin and Andrew Kowalczyk for their valuable comments on the manuscript. This work was supported by a Career Development Award from the Crohn's and Colitis Foundation of America.

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2008 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Ivanov, A.I. (2008). Pharmacological Inhibition of Endocytic Pathways: Is It Specific Enough to Be Useful?. In: Ivanov, A.I. (eds) Exocytosis and Endocytosis. Methods in Molecular Biology, vol 440. Humana Press. https://doi.org/10.1007/978-1-59745-178-9_2

Download citation

  • DOI: https://doi.org/10.1007/978-1-59745-178-9_2

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-865-2

  • Online ISBN: 978-1-59745-178-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics