title: Abnormal translocation of tyrosinase and tyrosinase-related protein 1 in cutaneous melanocytes of Hermansky-Pudlak Syndrome and in melanoma cells transfected with anti-sense HPS1 cDNA.
abstract: Hermansky-Pudlak syndrome is an autosomal recessive disorder characterized by oculocutaneous albinism, a bleeding disorder, and, in some patients, ceroid storage and progressive lung disease. Although Hermansky-Pudlak syndrome exhibits locus heterogeneity, most patients have mutations in the HPS1 gene. Melanocytes in the basal epithelial layer of skin from patients with different mutations in the HPS1 gene exhibited occasional large complexes containing dihydroxyphenylalanine-positive cisterna and 50 nm vesicles. To characterize the role of the HPS1 protein in cells, human HPS1 cDNA was transfected into pigmented SK-MEL-188 melanoma cells (M-188) in either the sense (S-188) or the antisense (A-188) orientation. Expression of the 79 kDa HPS1 protein (in M-188 and S-188 cells) or lack of expression (in A-188 cells) was confirmed by Western blotting using two HPS1-protein-specific polyclonal antibodies. Significant reduction in expression of HPS1 protein in A-188 cells resulted in a significant decrease in tyrosinase activity and melanin content compared with M-188 and S-188 cells using an intact cell assay for tyrosinase. In contrast, tyrosinase activities in cell lysates of M-188, S-188, and A-188 cells were not significantly different. Knockout of HPS1 protein expression in A-188 cells caused both tyrosinase and tyrosinase-related protein 1 to be localized to large granular complexes in the cell cytosol and dendrites. Electron microscope analysis of the A-188 cells revealed that absence of HPS1 protein resulted in the deposition of dihydroxyphenylalanine reaction products (i.e., tyrosinase) confined to large membrane-bound structures with limiting membranes. We conclude that lack of HPS1 protein expression results in mistranslocation of tyrosinase and tyrosinase-related protein 1 to large granular complexes rather than melanosomes, compromising melanin synthesis.
authors.affiliation: Department of Dermatology, University of Cincinnati, Cincinnati, Ohio 45267, USA.
authors.forename: R
authors.initials: R
authors.lastname: Sarangarajan
authors.forename: A
authors.initials: A
authors.lastname: Budev
authors.forename: Y
authors.initials: Y
authors.lastname: Zhao
authors.forename: W A
authors.initials: WA
authors.lastname: Gahl
authors.forename: R E
authors.initials: RE
authors.lastname: Boissy
mesh_terms: D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D005786:Gene Expression Regulation; D022861:Hermanski-Pudlak Syndrome / Q000235:genetics* / Q000473:pathology; D006801:Humans; D008544:Melanocytes / Q000473:pathology / Q000502:physiology; D008545:Melanoma / Q000235:genetics* / Q000473:pathology; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000235:genetics*; D014442:Monophenol Monooxygenase / Q000235:genetics*; D016376:Oligonucleotides, Antisense / Q000008:administration & dosage / Q000235:genetics; D010088:Oxidoreductases; D011506:Proteins / Q000235:genetics*; D012878:Skin Neoplasms / Q000235:genetics* / Q000473:pathology; D014162:Transfection; D014178:Translocation, Genetic
chemical_list: D018076:DNA, Complementary; C102160:HPS1 protein, human; C491131:Hps1 protein, mouse; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D016376:Oligonucleotides, Antisense; D011506:Proteins; D010088:Oxidoreductases; C489799:TYRP1 protein, human; C064056:tyrosinase-related protein-1; D014442:Monophenol Monooxygenase