Tyrosinase maturation and oligomerization in the endoplasmic reticulum require a melanocyte-specific factor.
Tyrosinase is a glycoprotein responsible for the synthesis of melanin in melanocytes. A large number of mutations have been identified in tyrosinase, with many leading to its misfolding, endoplasmic reticulum (ER) retention, and degradation. Here we describe the folding and maturation of human tyrosinase (TYR) using an in vitro translation system coupled with ER-derived microsomes or with semipermeabilized cells, as an intact ER source. TYR remained misfolded as determined by its sensitivity to trypsin digestion and its persistent interaction with the ER resident lectin chaperones calnexin and calreticulin when produced in ER-derived microsomes or nonmelanocytic semipermeabilized cells. However, when TYR was translocated into semipermeabilized melanocytes, chaperone interactions were transient, maturation progressed to a trypsin-resistant state, and a TYR homodimer was formed. The use of semipermeabilized mouse melanocytes defective for tyrosinase or other melanocyte-specific proteins as the ER source indicated that proper TYR maturation and oligomerization were greatly aided by the presence of wild type tyrosinase and tyrosinase-related protein 1. These findings suggested that oligomerization is a step in proper TYR maturation within the ER that requires melanocyte-specific factors.
Department of Biochemistry and Molecular Biology, Program in Molecular and Cellular Biology, University of Massachusetts, Amherst, Massachusetts 01003, USA.
Edwin
E
Francis
Ning
N
Wang
Hadas
H
Parag
Ruth
R
Halaban
Daniel N
DN
Hebert
D000818:Animals; D016466:CHO Cells; D037281:Calnexin / Q000378:metabolism; D037282:Calreticulin / Q000378:metabolism; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000494:pharmacology; D019281:Dimerization; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000201:enzymology* / Q000378:metabolism*; D037102:Lectins / Q000378:metabolism; D008544:Melanocytes / Q000378:metabolism*; D008562:Membrane Glycoproteins; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D014442:Monophenol Monooxygenase / Q000737:chemistry* / Q000378:metabolism*; D009154:Mutation; D010088:Oxidoreductases; D010179:Pancreas / Q000166:cytology / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011134:Polysaccharides / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D017510:Protein Folding; D021381:Protein Transport; D011506:Proteins / Q000378:metabolism; D011817:Rabbits; D013395:Sucrose / Q000378:metabolism / Q000494:pharmacology; D013997:Time Factors; D014158:Transcription, Genetic; D014357:Trypsin / Q000494:pharmacology
D037282:Calreticulin; D003432:Cross-Linking Reagents; D037102:Lectins; D008562:Membrane Glycoproteins; D011134:Polysaccharides; D011506:Proteins; D037281:Calnexin; D013395:Sucrose; D010088:Oxidoreductases; C489799:TYRP1 protein, human; C079621:Tyrp1 protein, mouse; C064056:tyrosinase-related protein-1; D014442:Monophenol Monooxygenase; D014357:Trypsin