Membrane trafficking regulates the activity of the human dopamine transporter.
The trafficking of synaptic proteins is unquestionably a major determinant of the properties of synaptic transmission. Here, we present a detailed analysis of the downregulation and intracellular trafficking of the cocaine- and amphetamine-sensitive dopamine transporter (DAT), a presynaptic plasma membrane protein responsible for the regulation of extracellular DA concentrations. Using PC12 cells stably transfected with human DAT cDNA, we observe that phorbol ester activation of protein kinase C (PKC) results in decreased transporter capacity and a parallel decrease in the amount of DAT on the cell surface that is attributable to intracellular transporter sequestration. After internalization, DAT diverges to the recycling, as opposed to the degradative, arm of the endocytic pathway. This study demonstrates, for the first time, DAT endocytosis, establishes the pathways through which DAT traffics both at steady state and in response to PKC activation, and suggests that DAT recycling is likely to occur.
Melikian
H E
HE
Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Buckley
K M
KM
D000818:Animals; D001692:Biological Transport; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002458:Cell Fractionation; D002462:Cell Membrane / Q000502:physiology*; D004298:Dopamine / Q000378:metabolism*; D050483:Dopamine Plasma Membrane Transport Proteins; D004705:Endocytosis; D011992:Endosomes / Q000502:physiology; D005786:Gene Expression Regulation; D006801:Humans; D007700:Kinetics; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D008959:Models, Neurological; D009419:Nerve Tissue Proteins; D016716:PC12 Cells; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D011990:Receptors, Transferrin / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D015398:Signal Transduction; D013572:Synaptic Vesicles / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002352:Carrier Proteins; D050483:Dopamine Plasma Membrane Transport Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; D011990:Receptors, Transferrin; D011994:Recombinant Proteins; C493214:SLC6A3 protein, human; C493216:Slc6a3 protein, rat; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D004298:Dopamine