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. 1984 Jan 1;98(1):108–115. doi: 10.1083/jcb.98.1.108

Evidence for both prelysosomal and lysosomal intermediates in endocytic pathways

PMCID: PMC2112994  PMID: 6707080

Abstract

Horseradish peroxidase (HRP), an enzyme internalized by fluid phase pinocytosis, has been used to study the process by which pinosome contents are delivered to lysosomes in Chinese hamster ovary cells. Pinosome contents were labeled by allowing cells to internalize HRP for 3-5 min. Following various chase times, cells were either processed for HRP and acid phosphatase (AcPase) cytochemistry or homogenized and fractionated in Percoll gradients. In Percoll gradients, pinosomes labeled by a 3-5 min HRP pulse behaved as a vesicle population more dense than plasma membrane and less dense than lysosomes. In pulse- chase experiments, internalized HRP was chased rapidly (3-6 min chase) to a density position intermediate between the "initial" pinocytic vesicle population and lysosomes. With longer chase periods, a progressive accumulation of HRP in more dense vesicles was observed. Correspondence between the HRP distribution and lysosomal marker distribution was reached after a approximately 1-h chase. By electron microscope cytochemistry of intact cells, the predominant class of HRP- positive vesicles after pulse uptakes or a 3-min chase period was characterized by a peripheral rim of reaction product and was AcPase negative. After 10-120-min chase periods, the predominant class of HRP- positive vesicles was characterized by luminal deposits and HRP activity was frequently observed in multivesicular bodies. HRP-positive vesicles after a 10- or 30-min chase were AcPase-positive. No HRP activity was detected in Golgi apparatus. Together these observations indicate that progressive processing of vesicular components of the vacuolar apparatus occurs at both a prelysosomal and lysosomal stage.

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Selected References

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  1. Adams C. J., Maurey K. M., Storrie B. Exocytosis of pinocytic contents by Chinese hamster ovary cells. J Cell Biol. 1982 Jun;93(3):632–637. doi: 10.1083/jcb.93.3.632. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. DE DUVE C., PRESSMAN B. C., GIANETTO R., WATTIAUX R., APPELMANS F. Tissue fractionation studies. 6. Intracellular distribution patterns of enzymes in rat-liver tissue. Biochem J. 1955 Aug;60(4):604–617. doi: 10.1042/bj0600604. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dunphy W. G., Fries E., Urbani L. J., Rothman J. E. Early and late functions associated with the Golgi apparatus reside in distinct compartments. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7453–7457. doi: 10.1073/pnas.78.12.7453. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Dunphy W. G., Rothman J. E. Compartmentation of asparagine-linked oligosaccharide processing in the Golgi apparatus. J Cell Biol. 1983 Jul;97(1):270–275. doi: 10.1083/jcb.97.1.270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Farquhar M. G., Palade G. E. The Golgi apparatus (complex)-(1954-1981)-from artifact to center stage. J Cell Biol. 1981 Dec;91(3 Pt 2):77s–103s. doi: 10.1083/jcb.91.3.77s. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. HUNTER W. M., GREENWOOD F. C. Preparation of iodine-131 labelled human growth hormone of high specific activity. Nature. 1962 May 5;194:495–496. doi: 10.1038/194495a0. [DOI] [PubMed] [Google Scholar]
  7. Hirsch J. G., Fedorko M. E. Ultrastructure of human leukocytes after simultaneous fixation with glutaraldehyde and osmium tetroxide and "postfixation" in uranyl acetate. J Cell Biol. 1968 Sep;38(3):615–627. doi: 10.1083/jcb.38.3.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. LaBadie J. H., Chapman K. P., Aronson N. N., Jr Glycoprotein catabolism in rat liver: Lysosomal digestion of iodinated asialo-fetuin. Biochem J. 1975 Nov;152(2):271–279. doi: 10.1042/bj1520271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Maxfield F. R. Weak bases and ionophores rapidly and reversibly raise the pH of endocytic vesicles in cultured mouse fibroblasts. J Cell Biol. 1982 Nov;95(2 Pt 1):676–681. doi: 10.1083/jcb.95.2.676. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Merion M., Poretz R. D. The resolution of two populations of lysosomal organelles containing endocytosed Wistaria floribunda agglutinin from murine fibroblasts. J Supramol Struct Cell Biochem. 1981;17(4):337–346. doi: 10.1002/jsscb.380170405. [DOI] [PubMed] [Google Scholar]
  11. Merion M., Sly W. S. The role of intermediate vesicles in the adsorptive endocytosis and transport of ligand to lysosomes by human fibroblasts. J Cell Biol. 1983 Mar;96(3):644–650. doi: 10.1083/jcb.96.3.644. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Pertoft H., Wärmegård B., Hök M. Heterogeneity of lysosomes originating from rat liver parenchymal cells. Metabolic relationship of subpopulations separated by density-gradient centrifugation. Biochem J. 1978 Jul 15;174(1):309–317. doi: 10.1042/bj1740309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Pool R. R., Jr, Maurey K. M., Storrie B. Characterization of pinocytic vesicles from CHO cells: resolution of pinosomes from lysosomes by analytical centrifugation. Cell Biol Int Rep. 1983 May;7(5):361–367. doi: 10.1016/0309-1651(83)90076-0. [DOI] [PubMed] [Google Scholar]
  14. REYNOLDS E. S. The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol. 1963 Apr;17:208–212. doi: 10.1083/jcb.17.1.208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Rome L. H., Garvin A. J., Allietta M. M., Neufeld E. F. Two species of lysosomal organelles in cultured human fibroblasts. Cell. 1979 May;17(1):143–153. doi: 10.1016/0092-8674(79)90302-7. [DOI] [PubMed] [Google Scholar]
  16. Roth J., Berger E. G. Immunocytochemical localization of galactosyltransferase in HeLa cells: codistribution with thiamine pyrophosphatase in trans-Golgi cisternae. J Cell Biol. 1982 Apr;93(1):223–229. doi: 10.1083/jcb.93.1.223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rothman J. E. The golgi apparatus: two organelles in tandem. Science. 1981 Sep 11;213(4513):1212–1219. doi: 10.1126/science.7268428. [DOI] [PubMed] [Google Scholar]
  18. Spurr A. R. A low-viscosity epoxy resin embedding medium for electron microscopy. J Ultrastruct Res. 1969 Jan;26(1):31–43. doi: 10.1016/s0022-5320(69)90033-1. [DOI] [PubMed] [Google Scholar]
  19. Stanners C. P., Eliceiri G. L., Green H. Two types of ribosome in mouse-hamster hybrid cells. Nat New Biol. 1971 Mar 10;230(10):52–54. doi: 10.1038/newbio230052a0. [DOI] [PubMed] [Google Scholar]
  20. Steinman R. M., Brodie S. E., Cohn Z. A. Membrane flow during pinocytosis. A stereologic analysis. J Cell Biol. 1976 Mar;68(3):665–687. doi: 10.1083/jcb.68.3.665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Steinman R. M., Cohn Z. A. The interaction of soluble horseradish peroxidase with mouse peritoneal macrophages in vitro. J Cell Biol. 1972 Oct;55(1):186–204. doi: 10.1083/jcb.55.1.186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Steinman R. M., Mellman I. S., Muller W. A., Cohn Z. A. Endocytosis and the recycling of plasma membrane. J Cell Biol. 1983 Jan;96(1):1–27. doi: 10.1083/jcb.96.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Steinman R. M., Silver J. M., Cohn Z. A. Pinocytosis in fibroblasts. Quantitative studies in vitro. J Cell Biol. 1974 Dec;63(3):949–969. doi: 10.1083/jcb.63.3.949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Storrie B., Dreesen T. D., Maurey K. M. Rapid cell surface appearance of endocytic membrane proteins in Chinese hamster ovary cells. Mol Cell Biol. 1981 Mar;1(3):261–268. doi: 10.1128/mcb.1.3.261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Storrie B., Edelson P. J. Distribution of concanavalin A in fibroblasts: direct endocytosis versus surface capping. Cell. 1977 Jul;11(3):707–717. doi: 10.1016/0092-8674(77)90087-3. [DOI] [PubMed] [Google Scholar]
  26. Tietze C., Schlesinger P., Stahl P. Mannose-specific endocytosis receptor of alveolar macrophages: demonstration of two functionally distinct intracellular pools of receptor and their roles in receptor recycling. J Cell Biol. 1982 Feb;92(2):417–424. doi: 10.1083/jcb.92.2.417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Tycko B., Maxfield F. R. Rapid acidification of endocytic vesicles containing alpha 2-macroglobulin. Cell. 1982 Mar;28(3):643–651. doi: 10.1016/0092-8674(82)90219-7. [DOI] [PubMed] [Google Scholar]

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