Iron uptake and storage are tightly regulated to guarantee sufficient iron for essential cellular processes and to prevent the production of damaging free radicals. The placenta is the entry site for iron, which is delivered to the developing embryo. Iron is taken up by syncytiotrophoblast cells and is transported unidirectionally from mother to fetus against a concentration gradient. Several iron transporters and regulators were recently characterized, including DMT1 and ferroportin/Ireg1 that transport iron through membranes, and HFE that regulates TfR-mediated iron uptake. In this study we demonstrate that in a differentiated choriocarcinoma cell line BeWo, HFE, and TfR are localized mainly in recycling endosomes and a small percentage of these complexes is observed in late endosomes with DMT1 while in trophoblast cells, the level of TfR is significantly lower and it is detected with HFE and DMT1 mainly in late endosomes. Most interestingly, TfR and HFE, as well as TfR and DMT1 interact in placental trophoblast cells. Based on previous and these data we suggest that the level of intracellular iron may regulate both TfR expression (on the post-transcriptional and the post-translational levels) and TfR trafficking/transcytosis in polarized cells. © 2005 Wiley-Liss, Inc.