Decrease of cholesterol in mouse melanoma causes secretion of lysosomal enzymes.
We examined the change in the subcellular distribution of a lysosomal enzyme, beta-glucuronidase (beta-G), caused by decreased cholesterol levels in mouse melanoma cells using an HMG-CoA reductase inhibitor, lovastatin and lipoprotein-deficient serum (LDS). There was a decrease in the cholesterol content of the cells and increased secretion of the mature form of beta-G located in lysosomes, as documented by Percoll density gradient fractionation, digitonin permeabilization and immunoprecipitation. Furthermore, another lysosomal enzyme, cathepsin H, was found to be released in the medium from cells treated with lovastatin. Both the precursor and mature forms of cathepsin H were detected in the medium of treated cells. Next, when cells were treated with LDS without lovastatin, concomitantly with the decrease in the levels of cholesterol and beta-G activity in the cells, beta-G activity in the medium increased. Also, the ratio of beta-G (3.2-fold) released in the medium from cells treated with Dulbecco's modified Eagle medium (D-MEM) containing lovastatin and LDS was higher than that (2.3-fold) on treatment with D-MEM containing LDS without lovastatin. From these results, it was suggested that the exocytosis of mature enzymes from lysosomes into the medium or mis-sorting of the lysosomal precursor forms to the medium was caused by the lovastatin- and/or LDS-induced decrease in the cholesterol content of the cells, although the mechanism of secretion by lysosomal enzymes differed somewhat.
Michihara
Akihiro
A
Faculty of Pharmacy and Pharmaceutical Sciences, Fukuyama University, Fukuyama, Hiroshima 729-0292, Japan. mitihara@fupharm.fukuyama-u.ac.jp
Toda
Ken
K
Suenobu
Michihisa
M
Akasaki
Kenji
K
Tsuji
Hiroshi
H
D000818:Animals; D000924:Anticholesteremic Agents / Q000494:pharmacology; D056655:Cathepsin H; D002403:Cathepsins / Q000032:analysis; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002784:Cholesterol / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D003546:Cysteine Endopeptidases / Q000032:analysis; D019362:Cytochrome P-450 CYP2B1 / Q000032:analysis; D004721:Endoplasmic Reticulum / Q000737:chemistry; D005966:Glucuronidase / Q000032:analysis* / Q000378:metabolism; D007700:Kinetics; D008074:Lipoproteins / Q000378:metabolism; D008148:Lovastatin / Q000494:pharmacology; D008247:Lysosomes / Q000737:chemistry / Q000201:enzymology*; D008297:Male; D008546:Melanoma, Experimental / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D051381:Rats; D017208:Rats, Wistar
D000924:Anticholesteremic Agents; D003470:Culture Media; D008074:Lipoproteins; D002784:Cholesterol; D008148:Lovastatin; D019362:Cytochrome P-450 CYP2B1; D005966:Glucuronidase; D002403:Cathepsins; D003546:Cysteine Endopeptidases; D056655:Cathepsin H; C534127:Ctsh protein, mouse; C534128:Ctsh protein, rat