Copper-stimulated endocytosis and degradation of the human copper transporter, hCtr1.
Copper uptake at the plasma membrane and subsequent delivery to copper-dependent enzymes is essential for many cellular processes, including mitochondrial oxidative phosphorylation, free radical detoxification, pigmentation, neurotransmitter synthesis, and iron metabolism. However, intracellular levels of this nutrient must be controlled because it is potentially toxic in excess concentrations. The hCtr1 protein functions in high affinity copper uptake at the plasma membrane of human cells. In this study, we demonstrate that levels of the hCtr1 protein at the plasma membrane of HEK293 cells were reduced when cells were exposed to elevated copper. This decrease in surface hCtr1 levels was associated with an increased rate of endocytosis, and low micromolar concentrations of copper were sufficient to stimulate this process. Inhibitors of clathrin-dependent endocytosis prevented the trafficking of hCtr1 from the plasma membrane, and newly internalized hCtr1 and transferrin were co-localized. Significantly, elevated copper concentrations also resulted in the degradation of the hCtr1 protein. Our findings suggest that hCtr1-mediated copper uptake into mammalian cells is regulated by a post-translational mechanism involving copper-stimulated endocytosis and degradation of the transporter.
Petris
Michael J
MJ
Department of Nutritional Sciences, University of Missouri, Columbia, Missouri 65211, USA. petrism@missouri.edu
Smith
Kathryn
K
Lee
Jaekwon
J
Thiele
Dennis J
DJ
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D027682:Cation Transport Proteins; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002746:Chlorpromazine / Q000494:pharmacology; D003300:Copper / Q000378:metabolism / Q000494:pharmacology*; D000079923:Copper Transporter 1; D006224:Cricetinae; D004705:Endocytosis; D005609:Free Radicals; D006801:Humans; D015151:Immunoblotting; D008565:Membrane Proteins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D018377:Neurotransmitter Agents / Q000378:metabolism; D012680:Sensitivity and Specificity; D013997:Time Factors; D014168:Transferrin / Q000378:metabolism
D027682:Cation Transport Proteins; D000079923:Copper Transporter 1; D005609:Free Radicals; D008565:Membrane Proteins; D018377:Neurotransmitter Agents; C490079:SLC31A1 protein, human; D014168:Transferrin; D003300:Copper; D002746:Chlorpromazine