Transient expression of high molecular weight, heat sensitive, trypsin-resistant form of tyrosinase in B-16 melanoma cells.
High molecular weight forms of tyrosinase have been found to be expressed during spontaneous remelanization of the amelanotic B-16 melanoma cells in culture as well as in melanotic tumors formed from amelanotic melanoma cells grown in C57BL/6J mice. Overnight extraction of the crude melanosomal fractions from such tumors and cultured melanoma cells reveal the presence of an additional DOPA-MBTH positive band well below the stacking gel. This band has been found to be alpha-PEP7 (antibody specific for tyrosinase) positive and alpha-PEP1 (antibody specific for TRP-1) negative on Western blot analysis. Heat treatment at 60 degrees C for 60 min results in the loss of this band and considerable loss of activity of the melanosomal extract. Trypsin treatment of these melanosomal extracts resulted in a minor change in the mobility of the high molecular weight band. SDS-PAGE under reduced conditions followed by Western blotting revealed that the high molecular weight band was lost and not detected by alpha-PEP7 or alpha-PEP1. These findings indicate that high molecular weight, heat sensitive and trypsin resistant forms of tyrosinase are transiently expressed in B-16 melanoma cells and tumors that are initiating remelanization following phenotypic drift towards the amelanotic state.
Sharma
A M
AM
Hindustan Lever Research Centre, Andheri (East), Chakala, Mumbai, India.
Jagadisan
B
B
Sheorain
V S
VS
Haldankar
V
V
Mojamdar
M
M
D000818:Animals; D004586:Electrophoresis; D005656:Fungal Proteins / Q000378:metabolism; D006358:Hot Temperature; D006801:Humans; D008543:Melanins / Q000096:biosynthesis*; D008546:Melanoma, Experimental / Q000201:enzymology; D051379:Mice; D008810:Mice, Inbred C57BL; D008970:Molecular Weight; D014442:Monophenol Monooxygenase / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D010858:Pigmentation; D014357:Trypsin / Q000494:pharmacology
D005656:Fungal Proteins; D008543:Melanins; D009363:Neoplasm Proteins; D014442:Monophenol Monooxygenase; D014357:Trypsin