Characterization of a melanosomal transport system in murine melanocytes mediating entry of the melanogenic substrate tyrosine.
In this study, we identify a transport system for tyrosine, the initial precursor of melanin synthesis, in the melanosomes of murine melanocytes. Melanosomes preloaded with tyrosine demonstrated countertransport of 10 microM [3H]tyrosine, indicating carrier-mediated transport. Melanosomal tyrosine transport was saturable, with an apparent Km for tyrosine transport of 54 microM and a maximal velocity of 15 pmol of tyrosine/unit of hexosaminidase/min. Transport was temperature-dependent (Ea = 7.5 kcal/mol) and showed stereospecificity for the l-isomer of tyrosine. Aromatic, neutral hydrophobic compounds (such as tryptophan and phenylalanine), as well as the small, bulky neutral amino acids (such as leucine, isoleucine, and methionine) competed for tyrosine transport. Tyrosine transport was inhibited by the classical system L analogue, 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid and by monoiodotyrosine, but not by cystine, lysine, glutamic acid, or 2-(methylamino)-isobutyric acid. Tyrosine transport showed no dependence on Na+ or K+, and did not require an acidic environment or the availability of free thiols. These results demonstrate the existence of a neutral amino acid carrier in murine melanocyte melanosomes which resembles the rat thyroid FRTL-5 lysosomal system h. This transport system is critical to the function of the melanosome since tyrosine is the essential substrate required for the synthesis of the pigment melanin.
Potterf
S B
SB
Laboratory of Cell Biology, NCI, National Institutes of Health, Bethesda, Maryland 20892, USA.
Muller
J
J
Bernardini
I
I
Tietze
F
F
Kobayashi
T
T
Hearing
V J
VJ
Gahl
W A
WA
D000596:Amino Acids / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D001692:Biological Transport; D002151:Calorimetry; D002460:Cell Line; D003594:Cytoplasmic Granules / Q000378:metabolism / Q000648:ultrastructure; D007700:Kinetics; D008544:Melanocytes / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D008810:Mice, Inbred C57BL; D008854:Microscopy, Electron; D014316:Tritium; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism*; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
D000596:Amino Acids; D014316:Tritium; D014443:Tyrosine; D001619:beta-N-Acetylhexosaminidases