Transfer of tyrosinase to melanosomes in Harding-Passey mouse melanoma.
The transfer of tyrosinase from microsomes into melanosomes, without passing through the cytosol in the Harding-Passey mouse melanoma cell, was confirmed by experiments carried out using a combination of radioisotope tracer techniques and immunoprecipitation. 3H-Labeled amino acid incorporation into tyrosinase present in the microsome, melanosome, and soluble fractions confirmed the precursor-product relationship of the enzyme in the microsome fraction and in the melanosome fraction. However, two forms of the enzyme, Ts1- and Ts2-tyrosinase, separated from the soluble fraction by polyacrylamide gel electrophoresis, were shown to play no role in the transfer since little or no incorporation of radioactivity into tyrosinase in this fraction was found. It is suggested that most tyrosinase observed in the soluble fraction does not leak from the melanosomes or the microsomes during homogenization, but comes from necrotic tumor cells. It appears that melanosomal and microsomal tyrosinase might be released from the membrane of necrotic cells modified by various degradation enzymes, considering the data on the recovery of tyrosinase from the soluble fraction, where one-third of total enzyme activity in the postnuclear fraction could not be increased, even when the postnuclear fraction of the tumor was further homogenized radically.
D000818:Animals; D000918:Antibody Specificity; D004156:Catechol Oxidase / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004792:Enzyme Precursors / Q000302:isolation & purification*; D007120:Immunochemistry; D008544:Melanocytes / Q000201:enzymology*; D008545:Melanoma / Q000201:enzymology*; D051379:Mice; D008861:Microsomes / Q000201:enzymology*; D014442:Monophenol Monooxygenase / Q000302:isolation & purification*; D009374:Neoplasms, Experimental / Q000201:enzymology; D012995:Solubility
D004792:Enzyme Precursors; D004156:Catechol Oxidase; D014442:Monophenol Monooxygenase