Stimulation of melanoblast pigmentation by 8-methoxypsoralen:the involvement of microphthalmia-associated transcription factor, the protein kinase a signal pathway, and proteasome-mediated degradation.
In this study, we used melb-a melanoblasts as a model to study mechanisms involved in stimulating melanocyte function in vitiliginous skin following exposure to 8-methoxypsoralen (8MOP). Melanin content and tyrosinase activity increased 3- and 7-fold, respectively, in melanoblasts treated with 8MOP for 6 d compared with untreated controls. The intracellular signal pathways involved in 8MOP-induced effects on melanoblasts were investigated, particularly the roles of protein kinase A and protein kinase C. Forskolin, a protein kinase A activator, mimicked and enhanced the 8MOP stimulation of melanoblast pigmentation whereas a protein kinase C activator, 1-oleoyl-2-acetylglycerol, had no effect, indicating that the protein kinase A pathway is involved rather than the protein kinase C pathway. Those observations were confirmed using inhibitors of the protein kinase A or protein kinase C pathways. Western blot and semiquantitative reverse transcriptase polymerase chain reaction were performed to assess the protein and mRNA expression levels of microphthalmia-associated transcription factor and tyrosinase in melanoblasts treated with 8MOP for 3 h, 6 h, 1 d, 3 d, or 6 d. Incubation with 8MOP stimulated microphthalmia-associated transcription factor protein and mRNA levels within 3 h, but, in contrast, tyrosinase mRNA and protein levels did not increase following 8MOP treatment until 1 d after treatment. The proteasome inhibitor lactacystin blocked the proteasome-mediated proteolysis of tyrosinase, and its effect on proteasomal function was enhanced by 8MOP. Taken together, these results show that 8MOP functions by initially stimulating levels of microphthalmia-associated transcription factor expression via activation of the protein kinase A pathway, which thereby stimulates tyrosinase expression and function and eventually leads to dramatic increases in melanin production by melanoblasts.
Lei
Tie Chi
TC
Pigment Cell Biology Section, Laboratory of Cell Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Virador
Victoria
V
Yasumoto
Ken-Ichi
K
Vieira
Wilfred D
WD
Toyofuku
Kazutomo
K
Hearing
Vincent J
VJ
D000818:Animals; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D008543:Melanins / Q000378:metabolism; D008544:Melanocytes / Q000166:cytology / Q000502:physiology*; D008730:Methoxsalen / Q000494:pharmacology*; D051379:Mice; D008810:Mice, Inbred C57BL; D051739:Microphthalmia-Associated Transcription Factor; D014442:Monophenol Monooxygenase / Q000235:genetics / Q000378:metabolism; D009097:Multienzyme Complexes / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction / Q000502:physiology*; D012880:Skin Pigmentation / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology
D004268:DNA-Binding Proteins; D008543:Melanins; D051739:Microphthalmia-Associated Transcription Factor; C497235:Mitf protein, mouse; D009097:Multienzyme Complexes; D012333:RNA, Messenger; D014157:Transcription Factors; D005576:Colforsin; D014442:Monophenol Monooxygenase; D017868:Cyclic AMP-Dependent Protein Kinases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D008730:Methoxsalen