Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism
Farnesyl diphosphate synthase (FPPS: EC2.5.1.10), a key enzyme in isoprenoid metabolic pathways, catalyzes the synthesis of farnesyl diphosphate (FPP) an intermediate in the biosynthesis of both sterol and non-sterol isoprenoid end products. The localization of FPPS to peroxisomes has been reported...
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| Format: | Article |
| Language: | English |
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Elsevier
1999-09-01
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| Series: | Journal of Lipid Research |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022227520334039 |
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| author | Sita D. Gupta Ryan S. Mehan Terese R. Tansey Hua-Tang Chen Gertrud Goping Israel Goldberg Ishaiahu Shechter |
| author_facet | Sita D. Gupta Ryan S. Mehan Terese R. Tansey Hua-Tang Chen Gertrud Goping Israel Goldberg Ishaiahu Shechter |
| author_sort | Sita D. Gupta |
| collection | DOAJ |
| description | Farnesyl diphosphate synthase (FPPS: EC2.5.1.10), a key enzyme in isoprenoid metabolic pathways, catalyzes the synthesis of farnesyl diphosphate (FPP) an intermediate in the biosynthesis of both sterol and non-sterol isoprenoid end products. The localization of FPPS to peroxisomes has been reported (Krisans, S. K., J. Ericsson, P. A. Edwards, and G. A. Keller. 1994. J. Biol. Chem. 269: 14165–14169). Using indirect immunofluorescence and immunoelectron microscopic techniques we show here that FPPS is localized predominantly in the peroxisomes of rat hepatoma H35 cells. However, the partial release of 60–70% of cellular FPPS activity is observed by selective permeabilization of these cells with digitonin. Under these conditions, lactate dehydrogenase, a cytosolic enzyme, is completely released whereas catalase, a known peroxisomal enzyme, is fully retained. Digitonin treatment of H35 cells differentially affects the release of other peroxisomal enzymes involved in isoprenoid metabolism. For instance, mevalonate kinase and phosphomevalonate kinase are almost totally released (95% and 91%, respectively), whereas 3-hydroxy-3-methylglutaryl-CoA reductase is fully retained. Indirect immunoflourescence studies indicate that FPPS is localized in peroxisomes of Chinese hamster ovary (CHO)-K1 cells but is dispersed in the cytosol of ZR-82 cells, a mutant that lacks peroxisomes. Unlike in H35 cells, FPPS is completely released upon digitonin permeabilization of CHO-K1 and ZR-82 cells. In contrast, under the same permeabilization conditions, catalase is fully retained in CHO-K1 cells but completely released from ZR-82 cells. These studies indicate that FPPS and other enzymes in the isoprenoid biosynthetic pathways, involved in the formation of FPP, are differentially associated with peroxisomes and may easily diffuse to the cytosol. Based on these observations, the significance and a possible regulatory model in the formation of isoprenoid end-products are discussed.—Gupta, S. D., R. S. Mehan, T. R. Tansey, H-T. Chen, G. Goping, I. Goldberg, and I. Shechter. Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism. J. Lipid Res. 1999. 40: 1572–1584. |
| first_indexed | 2024-12-14T02:36:08Z |
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| issn | 0022-2275 |
| language | English |
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| publishDate | 1999-09-01 |
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| spelling | doaj.art-39f6f95cc22d4c82ac9b3aa84d5024482022-12-21T23:20:08ZengElsevierJournal of Lipid Research0022-22751999-09-0140915721584Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolismSita D. Gupta0Ryan S. Mehan1Terese R. Tansey2Hua-Tang Chen3Gertrud Goping4Israel Goldberg5Ishaiahu Shechter6From the Department of Biochemistry and Molecular Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799From the Department of Biochemistry and Molecular Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799From the Department of Biochemistry and Molecular Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799From the Department of Biochemistry and Molecular Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799Department of Anatomy and Cell Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799From the Department of Biochemistry and Molecular Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799From the Department of Biochemistry and Molecular Biology, F. Edward Hébert School of Medicine, Uniformed Services University of the Health Sciences, Bethesda, MD 20814-4799; To whom correspondence should be addressed.Farnesyl diphosphate synthase (FPPS: EC2.5.1.10), a key enzyme in isoprenoid metabolic pathways, catalyzes the synthesis of farnesyl diphosphate (FPP) an intermediate in the biosynthesis of both sterol and non-sterol isoprenoid end products. The localization of FPPS to peroxisomes has been reported (Krisans, S. K., J. Ericsson, P. A. Edwards, and G. A. Keller. 1994. J. Biol. Chem. 269: 14165–14169). Using indirect immunofluorescence and immunoelectron microscopic techniques we show here that FPPS is localized predominantly in the peroxisomes of rat hepatoma H35 cells. However, the partial release of 60–70% of cellular FPPS activity is observed by selective permeabilization of these cells with digitonin. Under these conditions, lactate dehydrogenase, a cytosolic enzyme, is completely released whereas catalase, a known peroxisomal enzyme, is fully retained. Digitonin treatment of H35 cells differentially affects the release of other peroxisomal enzymes involved in isoprenoid metabolism. For instance, mevalonate kinase and phosphomevalonate kinase are almost totally released (95% and 91%, respectively), whereas 3-hydroxy-3-methylglutaryl-CoA reductase is fully retained. Indirect immunoflourescence studies indicate that FPPS is localized in peroxisomes of Chinese hamster ovary (CHO)-K1 cells but is dispersed in the cytosol of ZR-82 cells, a mutant that lacks peroxisomes. Unlike in H35 cells, FPPS is completely released upon digitonin permeabilization of CHO-K1 and ZR-82 cells. In contrast, under the same permeabilization conditions, catalase is fully retained in CHO-K1 cells but completely released from ZR-82 cells. These studies indicate that FPPS and other enzymes in the isoprenoid biosynthetic pathways, involved in the formation of FPP, are differentially associated with peroxisomes and may easily diffuse to the cytosol. Based on these observations, the significance and a possible regulatory model in the formation of isoprenoid end-products are discussed.—Gupta, S. D., R. S. Mehan, T. R. Tansey, H-T. Chen, G. Goping, I. Goldberg, and I. Shechter. Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism. J. Lipid Res. 1999. 40: 1572–1584.http://www.sciencedirect.com/science/article/pii/S0022227520334039peroxisomescholesterolisoprenoidsimmunofluorescencecellular permeabilizationfarnesyl diphosphate synthase |
| spellingShingle | Sita D. Gupta Ryan S. Mehan Terese R. Tansey Hua-Tang Chen Gertrud Goping Israel Goldberg Ishaiahu Shechter Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism Journal of Lipid Research peroxisomes cholesterol isoprenoids immunofluorescence cellular permeabilization farnesyl diphosphate synthase |
| title | Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism |
| title_full | Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism |
| title_fullStr | Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism |
| title_full_unstemmed | Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism |
| title_short | Differential binding of proteins to peroxisomes in rat hepatoma cells: unique association of enzymes involved in isoprenoid metabolism |
| title_sort | differential binding of proteins to peroxisomes in rat hepatoma cells unique association of enzymes involved in isoprenoid metabolism |
| topic | peroxisomes cholesterol isoprenoids immunofluorescence cellular permeabilization farnesyl diphosphate synthase |
| url | http://www.sciencedirect.com/science/article/pii/S0022227520334039 |
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