GENERATION OF THE SOLUBLE TRANSFERRIN RECEPTOR REQUIRES CYCLING THROUGH AN ENDOSOMAL COMPARTMENT

The transmembrane protein, transferrin receptor (TER), is found in a s oluble form in human serum and in the medium of cell lines grown in ti ssue culture, The soluble form is generated by proteolytic cleavage be tween Arg-100 and Leu-101. We used two mutant human TfRs expressed in Chinese hamster ovary (CHO) cells lacking endogenous transferrin recep tor to characterize the protease that cleaves the TfR and determine it s location in the cell, The T104D mutant TER lacks the O-linked carboh ydrate at position 104, and is more susceptible to proteolytic cleavag e at Arg-100 than the wildtype human TfR in these cells. We find that the protease is not a component of the serum in the growth medium, and it is not secreted by the cells, Cleavage does not occur during biosy nthesis of the TfR, and occurs after the TfR has reached the cell surf ace, Expression of the T104D TfR in a temperature-sensitive acidificat ion defective CHO cell line, G.7.1, shows that cleavage of the TfR is not dependent on acidification of endosomes, The C20A23 TfR is an endo cytosis deficient mutant lacking an internalization signal, This mutan t TfR, which is mainly localized to the cell surface, is cleaved less efficiently than the wild-type TfR, indicating that the protease is lo calized to an intracellular compartment.