The relationship between Na(+)/H(+) exchanger expression and tyrosinase activity in human melanocytes.
The activity of melanosome-associated tyrosinase in human melanocytes differs based on racial skin type. In melanocytes from Black skin, tyrosinase activity is high while in White melanocytes the activity of the enzyme is low. Recent studies suggest that low tyrosinase activity in White melanocytes may be due to an acidic pH environment within the melanosome. Because sodium/hydrogen (Na(+)/H(+)) exchangers (NHEs) are known to regulate intracellular pH, melanocytes were treated with NHE inhibitors to determine what effect this inhibition might have on tyrosinase activity. Treatment of Black melanocytes with ethyl-isopropyl amiloride (EIPA) caused a rapid dose-dependent inhibition of tyrosinase activity. This inhibition was not due to either direct enzyme inhibition or to a decrease in tyrosinase abundance. In contrast, treatment of White melanocytes with EIPA, cimetidine, or clonidine resulted in little inhibition of tyrosinase activity. Reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot analysis showed that both Black and White melanocytes expressed mRNA and protein for NHE-1, NHE-3, NHE-5, NHE-6, and NHE-7. Immunohistochemical analysis showed that NHE-7 and NHE-3 co-localized with the melanosomal protein, Tyrosinase Related Protein-1 (TRP-1). In addition, the vesicular proton pump, vesicular ATPase (V-ATPase), was found to be present in both White and Black melanosomes, indicating that organelles from both racial skin types are capable of being acidified. The results suggest that one or more NHEs may help regulate melanosome pH and tyrosinase activity in human melanocytes.
Smith
Dustin R
DR
Department of Biochemistry and Molecular Biology, The University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
Spaulding
Deborah T
DT
Glenn
Hayden M
HM
Fuller
Bryan B
BB
D017920:Antiporters / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D044383:Black People / Q000235:genetics; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007231:Infant, Newborn; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008543:Melanins / Q000378:metabolism*; D008544:Melanocytes / Q000378:metabolism*; D020460:Melanosomes / Q000378:metabolism; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014442:Monophenol Monooxygenase / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism; D020033:Protein Isoforms / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism; D044465:White People / Q000235:genetics
D017920:Antiporters; D004791:Enzyme Inhibitors; D008543:Melanins; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; C000613385:SLC9A3 protein, human; C429538:SLC9A7 protein, human; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D010088:Oxidoreductases; C064056:tyrosinase-related protein-1; D014442:Monophenol Monooxygenase; D025262:Vacuolar Proton-Translocating ATPases