The mouse brown (b) locus protein has dopachrome tautomerase activity and is located in lysosomes in transfected fibroblasts
| Autor: | Evald Rosengren, Anna Wittbjer, Alison J. Winder, Hans Rorsman |
|---|---|
| Rok vydání: | 1993 |
| Předmět: |
Tyrosinase
Molecular Sequence Data Golgi Apparatus Biology Mice Consensus Sequence medicine Animals Amino Acid Sequence Fibroblast Hair Color Isomerases Melanosome Glucuronidase chemistry.chemical_classification Melanins Expression vector Sequence Homology Amino Acid Monophenol Monooxygenase Cell Biology Transfection 3T3 Cells Fibroblasts Catalase Molecular biology Mice Mutant Strains Intramolecular Oxidoreductases Mice Inbred C57BL Enzyme medicine.anatomical_structure chemistry Peroxidases Concanavalin A Enzyme Induction biology.protein Lysosomes Dopachrome tautomerase Subcellular Fractions |
| Zdroj: | Journal of cell science. 106 |
| ISSN: | 0021-9533 |
| Popis: | Many genes mapping to pigmentation loci are involved in the regulation of melanin synthesis in the mouse. The brown (b) locus controls black/brown coat coloration, and its product has significant homology to the key melanogenic enzyme tyrosinase. This has led to suggestions that the b-protein is itself a melanogenic enzyme. In order to investigate its function, we have established lines of mouse fibroblasts stably expressing the b-protein by co-transfection of a b-protein expression vector and a plasmid conferring resistance to the antibiotic G418. The b-protein synthesised by these cells has the expected molecular mass of 75 kDa and reacts with three different anti-b-protein antibodies. We were unable to confirm previous reports that the b-protein has tyrosinase or catalase activity, but detected stereospecific dopachrome tautomerase activity in b-protein-expressing fibroblasts. This dopachrome tautomerase binds to Concanavalin A-Sepharose, and the major product of its action on L-dopachrome is 5,6-dihydroxyindole-2-carboxylic acid. Since this activity is not present in untransfected cells we conclude that the b-protein has dopachrome tautomerase activity. Fibroblasts do not contain melanosomes, the specialised organelles in which the b-protein is located in melanocytes. Nevertheless, indirect immunofluorescence localisation of the b-protein in transfected fibroblasts produces a distinctive pattern of intense juxtanuclear staining combined with punctate cytoplasmic staining. Double-labelling shows co-localisation of the b-protein with the late endosomal/lysosomal markers beta-glucuronidase and LAMP-1, both in transfected fibroblasts and in mouse melanoma cells. These findings are consistent with the hypothesis that melanosomes are closely related to lysosomes. |
| Databáze: | OpenAIRE |
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