Structural, functional, and tissue distribution analysis of human transferrin receptor-2 by murine monoclonal antibodies and a polyclonal antiserum.
Human transferrin receptor-2 (TFR-2) is a protein highly homologous to TFR-1/CD71 and is endowed with the ability to bind transferrin (TF) with low affinity. High levels of TFR-2 mRNA were found in the liver and in erythroid precursors. Mutations affecting the TFR-2 gene led to hemochromatosis type 3, a form of inherited iron overload. Several issues on distribution and function of the receptor were answered by raising a panel of 9 monoclonal antibodies specific for TFR-2 by immunizing mice with murine fibroblasts transfected with the human TFR-2 cDNA. A polyclonal antiserum was also produced in mice immunized with 3 peptides derived from the TFR-2 sequence, exploiting an innovative technique. The specificity of all the reagents produced was confirmed by reactivity with TFR-2(+) target cells and simultaneous negativity with TFR-1(+) cells. Western blot analyses showed a dominant chain of approximately 90 kDa in TFR-2 transfectants and HepG2 cell line. Analysis of distribution in normal tissues and in representative cell lines revealed that TFR-2 displays a restricted expression pattern--it is present at high levels in hepatocytes and in the epithelial cells of the small intestine, including the duodenal crypts. Exposure of human TFR-2(+) cells to TF-bound iron is followed by a significant up-regulation and relocalization of membrane TFR-2. The tissue distribution pattern, the behavior following exposure to iron-loaded TF, and the features of the disease resulting from TFR-2 inactivation support the hypothesis that TFR-2 contributes to body iron sensing.
Deaglio
Silvia
S
Laboratory of Immunogenetics, Department of Genetics, Biology, and Biochemistry, the Experimental Medicine Research Center, University of Turin Medical School, Italy.
Capobianco
Andrea
A
Calì
Angelita
A
Bellora
Francesca
F
Alberti
Federica
F
Righi
Luisella
L
Sapino
Anna
A
Camaschella
Clara
C
Malavasi
Fabio
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology; D000918:Antibody Specificity; D004727:Endothelium / Q000737:chemistry; D022781:Hepatocytes / Q000737:chemistry; D006801:Humans; D007106:Immune Sera / Q000276:immunology; D007150:Immunohistochemistry; D007422:Intestines / Q000166:cytology; D051379:Mice; D008807:Mice, Inbred BALB C; D011990:Receptors, Transferrin / Q000276:immunology* / Q000378:metabolism*; D014018:Tissue Distribution; D014168:Transferrin / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000187:drug effects
D000911:Antibodies, Monoclonal; D007106:Immune Sera; D011990:Receptors, Transferrin; C503640:TFR2 protein, human; C503641:TFR2 protein, mouse; D014168:Transferrin