Functional characterization of tyrosine transport in fibroblast cells from healthy controls.
Human fibroblast cells are an advantageous model to study the transport of amino acids across cell membranes, since one can control the environmental factors. A major problem in all earlier studies is the lack of precise and detailed knowledge regarding the expression and functionality of tyrosine transporters in human fibroblasts. This motivated us to perform a systematic functional characterization of the tyrosine transport in fibroblast cells with respect to the isoforms of system-L (LAT1, LAT2, LAT3, LAT4), which is the major transporter of tyrosine. Ten (n=10) fibroblast cell lines from healthy volunteers were included in the study. Uptake of L-[U-14C] tyrosine in fibroblasts was measured using the cluster tray method in the presence and absence of excess concentrations of various combinations of inhibitors. This study demonstrated that LAT1 is involved in 90% of total uptake of tyrosine and also around 51% of alanine. Not more than 10% can be accounted for by LAT2, LAT3 and LAT4 isoforms. LAT2 seems to be functionally weak in uptake of tyrosine while LAT3 and LAT4 contributed around 7%. 10% could be contributed by system-A (ATA2 isoform). Alanine consequently inhibited the tyrosine transport by up to 60%. Tyrosine transport through the LAT1 isoform has a higher affinity compared to system-L. In conclusion, the LAT1 isoform is the major transporter of tyrosine in human fibroblast cells. Competition between tyrosine and alanine for transport is shown to exist, probably between LAT1 and LAT2 isoforms. This study established fibroblast cells as a suitable experimental model for studying amino acid transport defects in humans.
Vumma
Ravi
R
Department of Clinical Medicine, Biomedicine, Orebro University, SE-701 82 Orebro, Sweden.
Wiesel
Frits-Axel
FA
Flyckt
Lena
L
Bjerkenstedt
Lars
L
Venizelos
Nikolaos
N
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000409:Alanine / Q000378:metabolism; D026923:Amino Acid Transport Systems, Basic / Q000378:metabolism; D001693:Biological Transport, Active / Q000502:physiology; D002250:Carbon Radioisotopes / Q000378:metabolism; D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism*; D006801:Humans; D027283:Large Neutral Amino Acid-Transporter 1 / Q000378:metabolism*; D020033:Protein Isoforms / Q000378:metabolism; D012016:Reference Values; D014443:Tyrosine / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D026923:Amino Acid Transport Systems, Basic; D002250:Carbon Radioisotopes; C419196:LAT2 protein, human; D027283:Large Neutral Amino Acid-Transporter 1; D020033:Protein Isoforms; C469951:SLC7A6 protein, human; D014443:Tyrosine; D000409:Alanine