Characteristics of the interaction between Hsc70 and the transferrin receptor in exosomes released during reticulocyte maturation

The transferrin receptor (TfR) of reticulocytes is released in vesicular fo rm (exosomes) during their maturation to erythrocytes, The heat shock cogna te 70-kDa protein (Hsc70) has been demonstrated to interact with the cytoso lic domain of the TfR and could thus trigger the receptor toward this secre tion pathway. We investigated the characteristics of the interaction betwee n Hsc70 and the TfR in exosomes with an in vitro binding assay using TfR im mobilized on Sepharose beads and purified Hsc70. The results show that Hsc7 0 binds to exosomal TfR with characteristics expected of a chaperone/peptid e interaction. We demonstrated that heat-denatured luciferase competed for in vitro binding, dependent on the nucleotide bound to Hsc70, and that this interaction activates the ATPase activity of Hsc70. Moreover, we used immu nosuppressive agents that interact with Hsc70, thus decreasing Hsc70 bindin g to TfR in our in vitro binding assay and enabling us to assess the role o f this interaction in vivo during reticulocyte maturation.