Regulation of transferrin-induced endocytosis by wild-type and C282Y-mutant HFE in transfected HeLa cells.
The hereditary hemochromatosis protein HFE is known to complex with the transferrin receptor; however, its function regarding endocytosis of transferrin is unclear. We performed patch-clamp capacitance measurements in transfected HeLa cells carrying wild-type or C282Y-mutant HFE cDNA under the control of a tetracycline-sensitive promoter. Whole cell experiments in cells with suppressed expression of wild-type HFE revealed a decrease in membrane capacitance, reflecting predominance of endocytosis in the presence of transferrin. Cells overexpressing C282Y-mutant HFE displayed less intense capacitance decreases, whereas no significant decrease was observed in cells overexpressing wild-type HFE. The formation of single endocytic vesicles in cells with suppressed expression of wild-type HFE was greatly increased in the presence of transferrin as revealed by cell-attached recordings. According to their calculated diameters, many of these vesicles corresponded to clathrin-coated vesicles. These results suggest that wild-type HFE negatively modulates the endocytic uptake of transferrin. This inhibitory effect is attenuated in cells expressing C282Y-mutant HFE. Time-resolved measurements of cell membrane capacitance provide a powerful tool to study transferrin-induced endocytosis in single cells.
Schwake
Lukas
L
Department of Gastroenterology, Faculty of Medicine, University of Heidelberg, D-69115 Heidelberg, Germany. Lukas_Schwake@gmx.net
Henkel
Andreas W
AW
Riedel
Hans D
HD
Schlenker
Thorsten
T
Both
Matthias
M
Migala
Andrea
A
Hadaschik
Boris
B
Henfling
Nataly
N
Stremmel
Wolfgang
W
D025802:Electric Capacitance; D006680:HLA Antigens / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006432:Hemochromatosis / Q000235:genetics*; D000071020:Hemochromatosis Protein; D015395:Histocompatibility Antigens Class I / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007501:Iron / Q000378:metabolism; D008565:Membrane Proteins; D018408:Patch-Clamp Techniques; D011990:Receptors, Transferrin / Q000378:metabolism; D022142:Secretory Vesicles / Q000378:metabolism; D014168:Transferrin / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism
C103673:HFE protein, human; D006680:HLA Antigens; D000071020:Hemochromatosis Protein; D015395:Histocompatibility Antigens Class I; D008565:Membrane Proteins; D011990:Receptors, Transferrin; D014168:Transferrin; D007501:Iron