Lysosomal enzyme secretion

Smith, R. J., and Ignarrr), L. J. (1975). Bioregulation of lysosomal enzyme. secretion from human neutrophils Roles of cyclic GMP and calcium in stimulus-secretion coupling. Proc. Natl. Acad. Sci. U.S.A. 72, 108-112. 

Historically, the important observation made by Hickman and Neufeld (1972) that I-cell fibroblasts are capable of endocytosing (the process by which cells take up macromolecules) lysosomal enzymes secreted by normal cells but that normal cells are incapable of internalizing the enzymes secreted by I-cell fibroblasts suggested that some kind of recognition marker for internalization of lysosomal enzymes was absent in I-cell fibroblasts. This recognition marker has subsequently been identified as mannose 6-phosphate. 

H4. Helminen, H. J., and Ericsson, J. L. E., On the mechanism of lysosomal enzyme secretion. Electron microscopic and histochemical studies on the epithelial cells of the rat s ventral prostate lobe. J. Ultrastruct. Res. 33, 528-549 (1970). 

The mechanisms of enzyme secretion and chemotaxis have obvious similarities. Thus, many chemotactic factors are also secretagogues for lysosomal enzyme secretion, e.g. fMLP [283,284]. Compounds causing secretion [285] and/or phagocytosis [286] increase the liberation of arachidonic acid from the phospholipids in a Ca -dependent manner [287]. Arachidonic acid in the presence of cytochalasin B causes enzyme release from rabbit peritoneal PMNL but not from human PMNL [152,288,290]. The acetylenic analog of arachidonic acid (5,8,11,14-eicosatetraenoic acid, ETYA) and nordihydroguiaiaretic acid (NDGA), both inhibitors of lipoxygenase, inhibit enzyme secretion induced by arachidonic acid [291,292] or chemotactic peptides and the complement factor C5a [289],... 

Joe B, Lokesh BR (1997) Effect of curcumin and capsaicin on arachidonic acid metabolism and lysosomal enzyme secretion by rat peritoneal macrophages. Lipids 32 1173-1180... 

Showell, H.J., Naccache, P.H., Walenga, R.W. et al. (1981). The effects of quercetin, 1-tosylamido-2-phenylethyl chloromethyl ketone, cytochalasin A and nordihydroguaiaretic acid on lysosomal enzyme secretion, arachidonic acid metabolism and Ca fluxes in rabbit neutrophils.. Journal of the Reticuloendothelial Society, 30, 167-181. 

Neutrophils represent an ideal system for studying osmotic effects on exocytosis. Stimulation of cytochalasin-B-treated neutrophils with the chemotactic peptide Jlf-formylmethionyl-leucyl-phenyl-alanine (FMLP) results in a rapid compound exocytosis up to 80% of lysosomal enzymes are released within 30 s (9-14). Secretion appears to be triggered by a rise in the level of cytosolic free calcium (15-18) promoted in part by entry of extracellular calcium through receptor-gated channels and in part by release of calcium that is sequestered or bound at some intracellular site (19-21). In this presentation, we augment our previously published data (22,23), which demonstrates that lysosomal enzyme release from neutrophils is inhibited under hyperosmotic conditions and that the rise in cytosolic calcium preceding secretion is inhibited as well. 

Cellular Effects of Complement Activation. C5a is chemotactic for neutrophils, activates their oxidative metabolism, and induces secretion of lysosomal enzymes from the granulocytes and macrophages. C5a may also induce the production of cytokines and prostaglandins (H19, S14). 

A successful tool in the early studies of metabolic pathways was blocking the pathway at some specific point. This could be done by the use of either mutants or inhibitors. Schekman et al have isolated a number of yeast mutants with blocks in their secretion pathway (Schekman, 1982). It is not yet known which proteins these mutations affect, but this is clearly a most promising approach for identifying those components involved in transport. In animal cells there are no cellular mutants with blocks in the intracellular transport of protein from the ER to the cell surface. There are, however, genetic diseases which affect the routing of lysosomal enzymes to the lysosomes (Neufeld et al, 1975 Sly and Fischer, 1982). For viruses it has been possible to isolate temperature-sensitive mutants in which a mutation in the viral glycoprotein arrests... 

Lysosomal enzymes are glycosylated and modified in a characteristic way. Most importantly, when they arrive in the Golgi apparatus, specific mannose residues in their oligosaccharide chains are phosphorylated. This phosphorylation is the critical event that removes them Corn the secretion pathway and directs them to lysosomes. Genetic defects affecting this phosphorylation produce I-ceU disease in which lysosomal enzymes are released into the ejctracellular space, and inclusion bodies accumulate in the cell, compromising its function. 

The answer is D. As this patient ages, a variety of skeletal defects and short stature that are consistent with a lysosomal storage disease (mucolipidosis), either I-cell disease or pseudo-Hurler polydystrophy, are developing. Both diseases arise from a deficiency of an enzyme involved in synthesis of the Man-6-P marker on lysosomal enzymes. Such misaddressed proteins are secreted rather than trafficked to the lysosomes. The degradative function of lysosomes is impaired as a result and the organelles tend to accumulate waste products (hence, the term storage disease ). It is these inclusion bodies or dense structures that would be visible by microscopic examination of the patient s cells in a biopsy specimen. 

A different conclusion was reached by Korchak et al. in studies of human PMNs. Blockers of the anion channel reduced the secretion of lysosomal enzymes in response to immune complexes and the ionophore for Ca, A 23187, but did not affect generation of O . Whether the differences in the results of the two studies are wholly attributable to the different species studied (bovine vs. human) or to other differences is not clear. 

Evidence has been presented that the concentration of cAMP is transiently increased in PMNs stimulated to form O2 by FMLP or Csa . Paradoxically, agents which increase the intracellular concentration of cAMP caused a dose dependent fall in the formation of O by PMNs which was elicited by FMLP. A rise in the concentration of cAMP within PMNs was also observed by Smolen after stimulation with several agents. The increase in the concentration of cAMP preceded the release of lysosomal enzymes and the formation of O but did not occur before the change in membrane potential. The increase in the concentration of cAMP was judged not to be sufficient for the elaboration of O2 or the secretion of enzymes from lysosomes because three maneuvers produced changes in cAMP but no subsequent response. However, whether the concentration of cAMP must rise in the normal chain of events leading from stimulus to formation of O is not clear. 

From study of peptides formed by partial hydrolysis of the 32P-labeled chymotrypsin, the sequence of amino acids surrounding the reactive serine was established and serine 195 was identified as the residue whose side chain hydroxyl group became phosphorylated. The same sequence Gly-Asp-Ser-Gly was soon discovered around reactive serine residues in trypsin, thrombin, elastase, and in the trypsin-like cocoonase used by silkmoths to escape from their cocoons.198 We know now that these are only a few of the enzymes in a very large family of serine proteases, most of which have related active site sequences.199 200 Among these are thrombin and other enzymes of the blood-clotting cascade (Fig. 12-17), proteases of lysosomes, and secreted proteases. 

Explain why fibroblasts from a patient with I-cell disease secrete lysosomal enzymes when grown in tissue culture. 

The secretion of thyroid hormones starts with endocy-tosis of the modified thyroglobulin, followed by fusion of the endocytotic vesicles with lysosomes. The lysosomal enzymes then degrade the thyroglobulin, liberating triiodothyronine and thyroxine into the circulation. Only about five molecules of T3 and T4 are generated from each molecule of thyroglobulin. Thyroid hormone secretion is stimulated by thyrotropin (TSH), a pituitary hormone that activates adenylate cyclase in its target cells. 

Cat B is an abundant and ubiquitously expressed cysteine peptidase of the papain family and makes up a major fraction of lysosomal enzymes that is capable of degrading components of the extracellular matrix in various diseases [30-32]. Cat B is also a prognostic marker for several types of cancer [33], and increased expression and secretion of cat B has been shown to be involved in the migration and invasion of various tumours [34—36], The precise role of cat B in solid tumours is not fully understood, but it has been proposed to participate, along with other cysteine cathepsins, in metastasis, angiogenesis, and tumour progression [37], Indeed, cat B inhibitors reduce both tumour cell motility and invasiveness in vitro [38], Recently, metal complexes based on rhenium, gold and palladium were shown to be effective inhibitors of cat B [39-44],... 

Biochemically, I-cell disease is characterized by excessive secretion of newly synthesized lysosomal enzymes into body fluids and concomitant loss of respective intracellular activities in fibroblasts. Shown in Table 17-1 are representative lysosomal enzyme activity levels in serum from patients with I-cell disease and those with the closely related disorder pseudo-Hurler poly dystrophy, indicating significantly increased levels of lysosomal enzyme activity. Germane to the biochemical diagnosis is the characteristic pattern of lysosomal enzyme deficiency in cultured fibroblasts, that is, an increase in the ratio of extracellular to intracellular enzyme activity (Table 17-2). It is interesting to note that not all lysosomal (i.e. intracellular)... 

Prenatal diagnosis of I-cell disease has been based on greatly reduced phosphotransferase activity (cf. Biochemical Perspectives section) and abnormal intracellular-extracellular distribution of lysosomal enzymes in cultured amni-otic fluid cells (Table 17-3).As indicated in Table 17-3, amniotic fluid cells secrete large amounts of lysosomal enzymes into the extracellular medium. Decreased levels of lysosomal enzymes in chorionic villi obtained by biopsy have also been observed in I-cell disease however, the characteristic secondary effect (i.e.,increased levels of lysosomal enzymes in the extracellular compartment) is only partially expressed or not expressed at all in chorionic villi, suggesting an alternative mechanism for the transport of lysosomal proteins. Although... 

The I-cell patient has proved invaluable for elucidation of the complex nature of intracellular packaging and sorting of lysosomal enzymes. The physiological importance of this signal-mediated pathway is evident in that fibroblasts from patients with I-cell disease and pseudo-Hurler polydystrophy secrete rather than target most of their lysosomal enzymes. Thus, the molecular theme of I-cell disease is that of faulty lysosomal targeting, the inability to transport (i.e., sort) lysosomal enzymes from their site of synthesis to the lysosome. 

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