Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1985 Jun 1;100(6):1894–1904. doi: 10.1083/jcb.100.6.1894

Effects of ammonia on processing and secretion of precursor and mature lysosomal enzyme from macrophages of normal and pale ear mice: evidence for two distinct pathways

PMCID: PMC2113587  PMID: 3922995

Abstract

Lysosomal enzymes have been shown to be synthesized as microsomal precursors, which are processed to mature enzymes located in lysosomes. We examined the effect of ammonium chloride on the intracellular processing and secretion of two lysosomal enzymes, beta-glucuronidase and beta-galactosidase, in mouse macrophages. This lysosomotropic drug caused extensive secretion of both precursor and mature enzyme forms within a few hours, as documented by pulse radiolabeling and molecular weight analysis. The normal intracellular route for processing and secretion of precursor enzyme was altered in treated cells. A small percentage of each precursor was delivered to the lysosomal organelle slowly. Most precursor forms traversed the Golgi apparatus, underwent further processing of carbohydrate moieties, and were then secreted in a manner similar to secretory proteins. The lag time for secretion of newly synthesized beta-galactosidase precursor was notably longer than that for the beta-glucuronidase precursor. The source of the secreted mature enzyme was the lysosomal organelle. Macrophages from the pale ear mutant were markedly deficient in secretion of mature lysosomal enzyme but secreted precursor forms normally. These results suggest that ammonia-treated macrophages contain two distinct intracellular pathways for secretion of lysosomal enzymes and that a specific block in the release of lysosomal contents occurs in the pale ear mutant.

Full Text

The Full Text of this article is available as a PDF (1.8 MB).

Selected References

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

  1. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  2. Brandt E. J., Elliott R. W., Swank R. T. Defective lysosomal enzyme secretion in kidneys of Chediak-Higashi (beige) mice. J Cell Biol. 1975 Dec;67(3):774–788. doi: 10.1083/jcb.67.3.774. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brown J. A., Swank R. T. Subcellular redistribution of newly synthesized macrophage lysosomal enzymes. Correlation between delivery to the lysosomes and maturation. J Biol Chem. 1983 Dec 25;258(24):15323–15328. [PubMed] [Google Scholar]
  4. Davies P., Bonney R. J. Secretory products of mononuclear phagocytes: a brief review. J Reticuloendothel Soc. 1979 Jul;26(1):37–47. [PubMed] [Google Scholar]
  5. Dean R. T., Jessup W., Roberts C. R. Effects of exogenous amines on mammalian cells, with particular reference to membrane flow. Biochem J. 1984 Jan 1;217(1):27–40. doi: 10.1042/bj2170027. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dimond R. L., Knecht D. A., Jordan K. B., Burns R. A., Livi G. P. Secretory mutants in the cellular slime mold Dictyostelium discoideum. Methods Enzymol. 1983;96:815–828. doi: 10.1016/s0076-6879(83)96069-x. [DOI] [PubMed] [Google Scholar]
  7. Erickson A. H., Blobel G. Carboxyl-terminal proteolytic processing during biosynthesis of the lysosomal enzymes beta-glucuronidase and cathepsin D. Biochemistry. 1983 Oct 25;22(22):5201–5205. doi: 10.1021/bi00291a021. [DOI] [PubMed] [Google Scholar]
  8. Fitting T., Kabat D. Evidence for a glycoprotein "signal" involved in transport between subcellular organelles. Two membrane glycoproteins encoded by murine leukemia virus reach the cell surface at different rates. J Biol Chem. 1982 Dec 10;257(23):14011–14017. [PubMed] [Google Scholar]
  9. Gibb S., Håkansson E. M., Lundin L. G., Shire J. G. Reduced pigmentation (rp), a new coat colour gene with effects on kidney lysosomal glycosidases in the mouse. Genet Res. 1981 Feb;37(1):95–103. doi: 10.1017/s0016672300020048. [DOI] [PubMed] [Google Scholar]
  10. Gonzalez-Noriega A., Grubb J. H., Talkad V., Sly W. S. Chloroquine inhibits lysosomal enzyme pinocytosis and enhances lysosomal enzyme secretion by impairing receptor recycling. J Cell Biol. 1980 Jun;85(3):839–852. doi: 10.1083/jcb.85.3.839. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gumbiner B., Kelly R. B. Two distinct intracellular pathways transport secretory and membrane glycoproteins to the surface of pituitary tumor cells. Cell. 1982 Jan;28(1):51–59. doi: 10.1016/0092-8674(82)90374-9. [DOI] [PubMed] [Google Scholar]
  12. Hasilik A., Neufeld E. F. Biosynthesis of lysosomal enzymes in fibroblasts. Synthesis as precursors of higher molecular weight. J Biol Chem. 1980 May 25;255(10):4937–4945. [PubMed] [Google Scholar]
  13. Hasilik A., Pohlmann R., von Figura K. Inhibition by cyanate of the processing of lysosomal enzymes. Biochem J. 1983 Mar 15;210(3):795–802. doi: 10.1042/bj2100795. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hasilik A., Von Figura K. Oligosaccharides in lysosomal enzymes. Distribution of high-mannose and complex oligosaccharides in cathepsin D and beta-hexosaminidase. Eur J Biochem. 1981 Dec;121(1):125–129. doi: 10.1111/j.1432-1033.1981.tb06440.x. [DOI] [PubMed] [Google Scholar]
  15. Imort M., Zühlsdorf M., Feige U., Hasilik A., von Figura K. Biosynthesis and transport of lysosomal enzymes in human monocytes and macrophages. Effects of ammonium chloride, zymosan and tunicamycin. Biochem J. 1983 Sep 15;214(3):671–678. doi: 10.1042/bj2140671. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jessup W., Leoni P., Bodmer J. L., Dean R. T. The effect of weak bases on lysosomal enzyme secretion by mononuclear phagocytes. Biochem Pharmacol. 1982 Aug 15;31(16):2657–2662. doi: 10.1016/0006-2952(82)90714-6. [DOI] [PubMed] [Google Scholar]
  17. Jessup W., Shirazi M. F., Dean R. T. Inhibition of some spontaneous secretory processes in macrophages and fibroblasts by ammonium chloride. Biochem Pharmacol. 1983 Sep 15;32(18):2703–2710. doi: 10.1016/0006-2952(83)90079-5. [DOI] [PubMed] [Google Scholar]
  18. Keeling P. J., Henson P. M. Lysosomal enzyme release from human monocytes in response to particulate stimuli. J Immunol. 1982 Feb;128(2):563–567. [PubMed] [Google Scholar]
  19. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  20. McCluer R. H., Williams M. A., Gross S. K., Meisler M. H. Testosterone effects on the induction and urinary excretion of mouse kidney glycosphingolipids associated with lysosomes. J Biol Chem. 1981 Dec 25;256(24):13112–13120. [PubMed] [Google Scholar]
  21. Meisler M. H. Synthesis and secretion of kidney beta-galactosidase in mutant le/le mice. J Biol Chem. 1978 May 10;253(9):3129–3134. [PubMed] [Google Scholar]
  22. Miller A. L., Kress B. C., Stein R., Kinnon C., Kern H., Schneider J. A., Harms E. Properties of N-acetyl-beta-D-hexosaminidase from isolated normal and I-cell lysosomes. J Biol Chem. 1981 Sep 10;256(17):9352–9362. [PubMed] [Google Scholar]
  23. Natowicz M., Baenziger J. U., Sly W. S. Structural studies of the phosphorylated high mannose-type oligosaccharides on human beta-glucuronidase. J Biol Chem. 1982 Apr 25;257(8):4412–4420. [PubMed] [Google Scholar]
  24. Novak E. K., Hui S. W., Swank R. T. Platelet storage pool deficiency in mouse pigment mutations associated with seven distinct genetic loci. Blood. 1984 Mar;63(3):536–544. [PubMed] [Google Scholar]
  25. Novak E. K., Hui S. W., Swank R. T. The mouse pale ear pigment mutant as a possible animal model for human platelet storage pool deficiency. Blood. 1981 Jan;57(1):38–43. [PubMed] [Google Scholar]
  26. Novak E. K., Swank R. T. Lysosomal dysfunctions associated with mutations at mouse pigment genes. Genetics. 1979 May;92(1):189–204. doi: 10.1093/genetics/92.1.189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Novak E. K., Wieland F., Jahreis G. P., Swank R. T. Altered secretion of kidney lysosomal enzymes in the mouse pigment mutants ruby-eye, ruby-eye-2-J, and maroon. Biochem Genet. 1980 Jun;18(5-6):549–561. doi: 10.1007/BF00484401. [DOI] [PubMed] [Google Scholar]
  28. Novick P., Ferro S., Schekman R. Order of events in the yeast secretory pathway. Cell. 1981 Aug;25(2):461–469. doi: 10.1016/0092-8674(81)90064-7. [DOI] [PubMed] [Google Scholar]
  29. Ohkuma S., Poole B. Cytoplasmic vacuolation of mouse peritoneal macrophages and the uptake into lysosomes of weakly basic substances. J Cell Biol. 1981 Sep;90(3):656–664. doi: 10.1083/jcb.90.3.656. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Palade G. Intracellular aspects of the process of protein synthesis. Science. 1975 Aug 1;189(4200):347–358. doi: 10.1126/science.1096303. [DOI] [PubMed] [Google Scholar]
  31. Pohlmann R., Krüger S., Hasilik A., von Figura K. Effect of monensin on intracellular transport and receptor-mediated endocytosis of lysosomal enzymes. Biochem J. 1984 Feb 1;217(3):649–658. doi: 10.1042/bj2170649. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. REEVES W. J., Jr, FIMOGNARI G. M. AN IMPROVED PROCEDURE FOR THE PREPARATION OF CRYSTALLINE LACTIC DEHYDROGENASE FROM HOG HEART. J Biol Chem. 1963 Dec;238:3853–3858. [PubMed] [Google Scholar]
  33. Riches D. W., Stanworth D. R. Evidence for a mechanism for the initiation of acid hydrolase secretion by macrophages that is functionally independent of alternative pathway complement activation. Biochem J. 1982 Mar 15;202(3):639–645. doi: 10.1042/bj2020639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Riches D. W., Stanworth D. R. Primary amines induce selective release of lysosomal enzymes from mouse macrophages. Biochem J. 1980 Jun 15;188(3):933–936. doi: 10.1042/bj1880933. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Riches D. W., Watkins J. L., Stanworth D. R. Biochemical differences in the mechanism of macrophage lysosomal exocytosis initiated by zymosan particles and weak bases. Biochem J. 1983 Jun 15;212(3):869–874. doi: 10.1042/bj2120869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Skudlarek M. D., Swank R. T. Biosynthesis of two lysosomal enzymes in macrophages. Evidence for a precursor of beta-galactosidase. J Biol Chem. 1979 Oct 25;254(20):9939–9942. [PubMed] [Google Scholar]
  37. Skudlarek M. D., Swank R. T. Turnover of two lysosomal enzymes in macrophages. J Biol Chem. 1981 Oct 10;256(19):10137–10144. [PubMed] [Google Scholar]
  38. Strous G. J., Lodish H. F. Intracellular transport of secretory and membrane proteins in hepatoma cells infected by vesicular stomatitis virus. Cell. 1980 Dec;22(3):709–717. doi: 10.1016/0092-8674(80)90547-4. [DOI] [PubMed] [Google Scholar]
  39. Swank R. T., Brandt E. J. Turnover of kidney beta-glucuronidase in normal and Chédiak-Higashi (beige) mice. Am J Pathol. 1978 Sep;92(3):755–772. [PMC free article] [PubMed] [Google Scholar]
  40. 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]
  41. Unanue E. R. Secretory function of mononuclear phagocytes: a review. Am J Pathol. 1976 May;83(2):396–418. [PMC free article] [PubMed] [Google Scholar]
  42. Varki A., Kornfeld S. The spectrum of anionic oligosaccharides released by endo-beta-N-acetylglucosaminidase H from glycoproteins. Structural studies and interactions with the phosphomannosyl receptor. J Biol Chem. 1983 Mar 10;258(5):2808–2818. [PubMed] [Google Scholar]
  43. Vladutiu G. D. The effect of chloroquine on the distribution of newly synthesized and old beta-hexosaminidase in fibroblasts. Biochem J. 1982 Dec 15;208(3):559–566. doi: 10.1042/bj2080559. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. de Duve C., de Barsy T., Poole B., Trouet A., Tulkens P., Van Hoof F. Commentary. Lysosomotropic agents. Biochem Pharmacol. 1974 Sep 15;23(18):2495–2531. doi: 10.1016/0006-2952(74)90174-9. [DOI] [PubMed] [Google Scholar]

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

RESOURCES