Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1994 Mar 1;124(5):689–703. doi: 10.1083/jcb.124.5.689

The coated pit and macropinocytic pathways serve distinct endosome populations

PMCID: PMC2119947  PMID: 8120092

Abstract

Clathrin-coated vesicle endocytosis and macropinocytosis are distinct endocytic pathways demonstrable in several cell types including human epidermoid A431 cells (West, M.A., M.S. Bretscher, and C. Watts. 1989. J. Cell Biol. 109:2731-2739). Here we analyze the extent of mixing of macropinocytic endosome (macropinosome) content with that of conventional endosomes served by coated vesicle endocytosis. Using laser scanning confocal fluorescence microscopy we detected very little delivery of macropinosome content to either early or late endosomes- lysosomes as defined by labeling with transferrin or with LDL. Mixing of the contents of the macropinosomes and conventional endosomes was not induced by the addition of brefeldin A. Moreover, the morphology of macropinosomes was not grossly altered in the presence of brefeldin A, whilst in the same cells there were dramatic tubulation effects on conventional endosomes as reported by others. Although refractory to fusion with conventional endosomes, macropinosomes were nonetheless dynamic structures which sometimes exhibited vesiculo-tubular morphology in living cells and were capable of fusing with each other. We suggest that different endocytic mechanisms can give rise to distinct endosome populations.

Full Text

The Full Text of this article is available as a PDF (6.8 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bar-Sagi D., Feramisco J. R. Induction of membrane ruffling and fluid-phase pinocytosis in quiescent fibroblasts by ras proteins. Science. 1986 Sep 5;233(4768):1061–1068. doi: 10.1126/science.3090687. [DOI] [PubMed] [Google Scholar]
  2. Bomsel M., Prydz K., Parton R. G., Gruenberg J., Simons K. Endocytosis in filter-grown Madin-Darby canine kidney cells. J Cell Biol. 1989 Dec;109(6 Pt 2):3243–3258. doi: 10.1083/jcb.109.6.3243. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bucci C., Parton R. G., Mather I. H., Stunnenberg H., Simons K., Hoflack B., Zerial M. The small GTPase rab5 functions as a regulatory factor in the early endocytic pathway. Cell. 1992 Sep 4;70(5):715–728. doi: 10.1016/0092-8674(92)90306-w. [DOI] [PubMed] [Google Scholar]
  4. Chavrier P., Gorvel J. P., Stelzer E., Simons K., Gruenberg J., Zerial M. Hypervariable C-terminal domain of rab proteins acts as a targeting signal. Nature. 1991 Oct 24;353(6346):769–772. doi: 10.1038/353769a0. [DOI] [PubMed] [Google Scholar]
  5. Colombo M. I., Gonzalo S., Weidman P., Stahl P. Characterization of trypsin-sensitive factor(s) required for endosome-endosome fusion. J Biol Chem. 1991 Dec 5;266(34):23438–23445. [PubMed] [Google Scholar]
  6. Damke H., Klumperman J., von Figura K., Braulke T. Effects of brefeldin A on the endocytic route. Redistribution of mannose 6-phosphate/insulin-like growth factor II receptors to the cell surface. J Biol Chem. 1991 Dec 25;266(36):24829–24833. [PubMed] [Google Scholar]
  7. Dowrick P., Kenworthy P., McCann B., Warn R. Circular ruffle formation and closure lead to macropinocytosis in hepatocyte growth factor/scatter factor-treated cells. Eur J Cell Biol. 1993 Jun;61(1):44–53. [PubMed] [Google Scholar]
  8. Dupree P., Parton R. G., Raposo G., Kurzchalia T. V., Simons K. Caveolae and sorting in the trans-Golgi network of epithelial cells. EMBO J. 1993 Apr;12(4):1597–1605. doi: 10.1002/j.1460-2075.1993.tb05804.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. FAWCETT D. W. SURFACE SPECIALIZATIONS OF ABSORBING CELLS. J Histochem Cytochem. 1965 Feb;13:75–91. doi: 10.1177/13.2.75. [DOI] [PubMed] [Google Scholar]
  10. Falkow S., Isberg R. R., Portnoy D. A. The interaction of bacteria with mammalian cells. Annu Rev Cell Biol. 1992;8:333–363. doi: 10.1146/annurev.cb.08.110192.002001. [DOI] [PubMed] [Google Scholar]
  11. Galán J. E., Pace J., Hayman M. J. Involvement of the epidermal growth factor receptor in the invasion of cultured mammalian cells by Salmonella typhimurium. Nature. 1992 Jun 18;357(6379):588–589. doi: 10.1038/357588a0. [DOI] [PubMed] [Google Scholar]
  12. Goldstein J. L., Brown M. S., Anderson R. G., Russell D. W., Schneider W. J. Receptor-mediated endocytosis: concepts emerging from the LDL receptor system. Annu Rev Cell Biol. 1985;1:1–39. doi: 10.1146/annurev.cb.01.110185.000245. [DOI] [PubMed] [Google Scholar]
  13. Gorvel J. P., Chavrier P., Zerial M., Gruenberg J. rab5 controls early endosome fusion in vitro. Cell. 1991 Mar 8;64(5):915–925. doi: 10.1016/0092-8674(91)90316-q. [DOI] [PubMed] [Google Scholar]
  14. Greenberg S., Chang P., Silverstein S. C. Tyrosine phosphorylation is required for Fc receptor-mediated phagocytosis in mouse macrophages. J Exp Med. 1993 Feb 1;177(2):529–534. doi: 10.1084/jem.177.2.529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gruenberg J., Howell K. E. Membrane traffic in endocytosis: insights from cell-free assays. Annu Rev Cell Biol. 1989;5:453–481. doi: 10.1146/annurev.cb.05.110189.002321. [DOI] [PubMed] [Google Scholar]
  16. HAVEL R. J., EDER H. A., BRAGDON J. H. The distribution and chemical composition of ultracentrifugally separated lipoproteins in human serum. J Clin Invest. 1955 Sep;34(9):1345–1353. doi: 10.1172/JCI103182. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Haigler H. T., McKanna J. A., Cohen S. Rapid stimulation of pinocytosis in human carcinoma cells A-431 by epidermal growth factor. J Cell Biol. 1979 Oct;83(1):82–90. doi: 10.1083/jcb.83.1.82. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hansen S. H., Sandvig K., van Deurs B. Molecules internalized by clathrin-independent endocytosis are delivered to endosomes containing transferrin receptors. J Cell Biol. 1993 Oct;123(1):89–97. doi: 10.1083/jcb.123.1.89. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hansen S. H., Sandvig K., van Deurs B. The preendosomal compartment comprises distinct coated and noncoated endocytic vesicle populations. J Cell Biol. 1991 May;113(4):731–741. doi: 10.1083/jcb.113.4.731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hopkins C. R., Gibson A., Shipman M., Miller K. Movement of internalized ligand-receptor complexes along a continuous endosomal reticulum. Nature. 1990 Jul 26;346(6282):335–339. doi: 10.1038/346335a0. [DOI] [PubMed] [Google Scholar]
  21. Hubbard A. L. Endocytosis. Curr Opin Cell Biol. 1989 Aug;1(4):675–683. doi: 10.1016/0955-0674(89)90033-1. [DOI] [PubMed] [Google Scholar]
  22. Hughson E. J., Hopkins C. R. Endocytic pathways in polarized Caco-2 cells: identification of an endosomal compartment accessible from both apical and basolateral surfaces. J Cell Biol. 1990 Feb;110(2):337–348. doi: 10.1083/jcb.110.2.337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hunziker W., Whitney J. A., Mellman I. Selective inhibition of transcytosis by brefeldin A in MDCK cells. Cell. 1991 Nov 1;67(3):617–627. doi: 10.1016/0092-8674(91)90535-7. [DOI] [PubMed] [Google Scholar]
  24. Joiner K. A., Fuhrman S. A., Miettinen H. M., Kasper L. H., Mellman I. Toxoplasma gondii: fusion competence of parasitophorous vacuoles in Fc receptor-transfected fibroblasts. Science. 1990 Aug 10;249(4969):641–646. doi: 10.1126/science.2200126. [DOI] [PubMed] [Google Scholar]
  25. Killisch I., Steinlein P., Römisch K., Hollinshead R., Beug H., Griffiths G. Characterization of early and late endocytic compartments of the transferrin cycle. Transferrin receptor antibody blocks erythroid differentiation by trapping the receptor in the early endosome. J Cell Sci. 1992 Sep;103(Pt 1):211–232. doi: 10.1242/jcs.103.1.211. [DOI] [PubMed] [Google Scholar]
  26. Klausner R. D., Donaldson J. G., Lippincott-Schwartz J. Brefeldin A: insights into the control of membrane traffic and organelle structure. J Cell Biol. 1992 Mar;116(5):1071–1080. doi: 10.1083/jcb.116.5.1071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Knapp P. E., Swanson J. A. Plasticity of the tubular lysosomal compartment in macrophages. J Cell Sci. 1990 Mar;95(Pt 3):433–439. doi: 10.1242/jcs.95.3.433. [DOI] [PubMed] [Google Scholar]
  28. Lippincott-Schwartz J., Yuan L., Tipper C., Amherdt M., Orci L., Klausner R. D. Brefeldin A's effects on endosomes, lysosomes, and the TGN suggest a general mechanism for regulating organelle structure and membrane traffic. Cell. 1991 Nov 1;67(3):601–616. doi: 10.1016/0092-8674(91)90534-6. [DOI] [PubMed] [Google Scholar]
  29. Montesano R., Roth J., Robert A., Orci L. Non-coated membrane invaginations are involved in binding and internalization of cholera and tetanus toxins. Nature. 1982 Apr 15;296(5858):651–653. doi: 10.1038/296651a0. [DOI] [PubMed] [Google Scholar]
  30. Mueller S. C., Hubbard A. L. Receptor-mediated endocytosis of asialoglycoproteins by rat hepatocytes: receptor-positive and receptor-negative endosomes. J Cell Biol. 1986 Mar;102(3):932–942. doi: 10.1083/jcb.102.3.932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Prydz K., Hansen S. H., Sandvig K., van Deurs B. Effects of brefeldin A on endocytosis, transcytosis and transport to the Golgi complex in polarized MDCK cells. J Cell Biol. 1992 Oct;119(2):259–272. doi: 10.1083/jcb.119.2.259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Racoosin E. L., Swanson J. A. M-CSF-induced macropinocytosis increases solute endocytosis but not receptor-mediated endocytosis in mouse macrophages. J Cell Sci. 1992 Aug;102(Pt 4):867–880. doi: 10.1242/jcs.102.4.867. [DOI] [PubMed] [Google Scholar]
  33. Racoosin E. L., Swanson J. A. Macrophage colony-stimulating factor (rM-CSF) stimulates pinocytosis in bone marrow-derived macrophages. J Exp Med. 1989 Nov 1;170(5):1635–1648. doi: 10.1084/jem.170.5.1635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Racoosin E. L., Swanson J. A. Macropinosome maturation and fusion with tubular lysosomes in macrophages. J Cell Biol. 1993 Jun;121(5):1011–1020. doi: 10.1083/jcb.121.5.1011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Ridley A. J., Paterson H. F., Johnston C. L., Diekmann D., Hall A. The small GTP-binding protein rac regulates growth factor-induced membrane ruffling. Cell. 1992 Aug 7;70(3):401–410. doi: 10.1016/0092-8674(92)90164-8. [DOI] [PubMed] [Google Scholar]
  36. Rothberg K. G., Heuser J. E., Donzell W. C., Ying Y. S., Glenney J. R., Anderson R. G. Caveolin, a protein component of caveolae membrane coats. Cell. 1992 Feb 21;68(4):673–682. doi: 10.1016/0092-8674(92)90143-z. [DOI] [PubMed] [Google Scholar]
  37. Rothberg K. G., Ying Y. S., Kamen B. A., Anderson R. G. Cholesterol controls the clustering of the glycophospholipid-anchored membrane receptor for 5-methyltetrahydrofolate. J Cell Biol. 1990 Dec;111(6 Pt 2):2931–2938. doi: 10.1083/jcb.111.6.2931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Rothberg K. G., Ying Y. S., Kolhouse J. F., Kamen B. A., Anderson R. G. The glycophospholipid-linked folate receptor internalizes folate without entering the clathrin-coated pit endocytic pathway. J Cell Biol. 1990 Mar;110(3):637–649. doi: 10.1083/jcb.110.3.637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Salzman N. H., Maxfield F. R. Fusion accessibility of endocytic compartments along the recycling and lysosomal endocytic pathways in intact cells. J Cell Biol. 1989 Nov;109(5):2097–2104. doi: 10.1083/jcb.109.5.2097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Salzman N. H., Maxfield F. R. Intracellular fusion of sequentially formed endocytic compartments. J Cell Biol. 1988 Apr;106(4):1083–1091. doi: 10.1083/jcb.106.4.1083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sandvig K., van Deurs B. Selective modulation of the endocytic uptake of ricin and fluid phase markers without alteration in transferrin endocytosis. J Biol Chem. 1990 Apr 15;265(11):6382–6388. [PubMed] [Google Scholar]
  42. Schmid S. L., Fuchs R., Male P., Mellman I. Two distinct subpopulations of endosomes involved in membrane recycling and transport to lysosomes. Cell. 1988 Jan 15;52(1):73–83. doi: 10.1016/0092-8674(88)90532-6. [DOI] [PubMed] [Google Scholar]
  43. Smythe E., Warren G. The mechanism of receptor-mediated endocytosis. Eur J Biochem. 1991 Dec 18;202(3):689–699. doi: 10.1111/j.1432-1033.1991.tb16424.x. [DOI] [PubMed] [Google Scholar]
  44. Swanson J. A. Phorbol esters stimulate macropinocytosis and solute flow through macrophages. J Cell Sci. 1989 Sep;94(Pt 1):135–142. doi: 10.1242/jcs.94.1.135. [DOI] [PubMed] [Google Scholar]
  45. Söllner T., Whiteheart S. W., Brunner M., Erdjument-Bromage H., Geromanos S., Tempst P., Rothman J. E. SNAP receptors implicated in vesicle targeting and fusion. Nature. 1993 Mar 25;362(6418):318–324. doi: 10.1038/362318a0. [DOI] [PubMed] [Google Scholar]
  46. Tooze J., Hollinshead M. In AtT20 and HeLa cells brefeldin A induces the fusion of tubular endosomes and changes their distribution and some of their endocytic properties. J Cell Biol. 1992 Aug;118(4):813–830. doi: 10.1083/jcb.118.4.813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Tooze J., Hollinshead M. Tubular early endosomal networks in AtT20 and other cells. J Cell Biol. 1991 Nov;115(3):635–653. doi: 10.1083/jcb.115.3.635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Watts C., Marsh M. Endocytosis: what goes in and how? J Cell Sci. 1992 Sep;103(Pt 1):1–8. doi: 10.1242/jcs.103.1.1a. [DOI] [PubMed] [Google Scholar]
  49. West M. A., Bretscher M. S., Watts C. Distinct endocytotic pathways in epidermal growth factor-stimulated human carcinoma A431 cells. J Cell Biol. 1989 Dec;109(6 Pt 1):2731–2739. doi: 10.1083/jcb.109.6.2731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Wood S. A., Park J. E., Brown W. J. Brefeldin A causes a microtubule-mediated fusion of the trans-Golgi network and early endosomes. Cell. 1991 Nov 1;67(3):591–600. doi: 10.1016/0092-8674(91)90533-5. [DOI] [PubMed] [Google Scholar]
  51. Woodman P. G., Mundy D. I., Cohen P., Warren G. Cell-free fusion of endocytic vesicles is regulated by phosphorylation. J Cell Biol. 1992 Jan;116(2):331–338. doi: 10.1083/jcb.116.2.331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Woodman P. G., Warren G. Fusion between vesicles from the pathway of receptor-mediated endocytosis in a cell-free system. Eur J Biochem. 1988 Apr 5;173(1):101–108. doi: 10.1111/j.1432-1033.1988.tb13972.x. [DOI] [PubMed] [Google Scholar]
  53. van Deurs B., Holm P. K., Sandvig K., Hansen S. H. Are caveolae involved in clathrin-independent endocytosis? Trends Cell Biol. 1993 Aug;3(8):249–251. doi: 10.1016/0962-8924(93)90045-3. [DOI] [PubMed] [Google Scholar]
  54. van Deurs B., Petersen O. W., Olsnes S., Sandvig K. The ways of endocytosis. Int Rev Cytol. 1989;117:131–177. doi: 10.1016/s0074-7696(08)61336-4. [DOI] [PubMed] [Google Scholar]
  55. van der Sluijs P., Hull M., Webster P., Mâle P., Goud B., Mellman I. The small GTP-binding protein rab4 controls an early sorting event on the endocytic pathway. Cell. 1992 Sep 4;70(5):729–740. doi: 10.1016/0092-8674(92)90307-x. [DOI] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES