Transferrin receptor 2 (TfR2) is a newly described protein with sequence similarity to the classical transferrin receptor (TfR). The function of TfR2 is unknown. In contrast to TfR, TfR2 is found almost exclusively in hepatocytes and early erythroid progenitor cells. Mutated forms of TfR2 are implicated in a form of hereditary hemochromatosis which results in iron overload in the liver. The liver is a major iron-processing organ in the body and disturbances in iron homeostasis in this organ could affect iron absorption by the intestines. On the basis of the limited information concerning this newly identified participant in iron homeostasis, the following hypotheses as to how TfR2 could plausibly participate in iron homeostasis by any or all of three different mechanisms will be tested. 1) TfR2 could regulate the efflux of iron out of the hepatocyte by facilitating the secretion of iron-loaded transferrin (Tf). Cell lines expressing TfR will be characterized for the rates of Tf synthesis, processing and iron-loading. 2) The binding of Tf to TfR2 or the uptake of iron via TfR2 could alter the secretion of iron-status signals from the hepatocytes. Alterations in the proteins secreted by hepatocytes will be examined by two-dimensional electrophoresis and mass spectrometry. Binding partners of TfR2 will be examined by immunoprecipitation and yeast two-hybrid studies to determine possible signaling mechanisms. 3) TfR2 could be located in the body in other locations that were not detected by northern analysis. Immunohistological staining of tissues with a monoclonal antibody generated against TfR2 will be used to examine specific cell types expressing TfR2. The long term goal of this research is to understand how the body regulates iron homeostasis and the mechanisms by which mutations in the key proteins result in disease.