Association of human transferrin receptor with GABARAP.

A yeast two-hybrid screen identified a specific interaction between the cytoplasmic domain of transferrin receptor (TfR) and GABARAP, a 14 kDa protein that binds to the gamma2 subunit of neuronal GABA(A) receptors. The specificity of the TfR-GABARAP interaction was demonstrated by in vitro binding assays with purified proteins and by co-immunoprecipitation of GABARAP with endogenous TfR from HeLa cell lysates. Replacement of the YTRF internalization motif with ATRA within the cytoplasmic domain of TfR reduced interaction with GABARAP in the yeast two-hybrid screen and in vitro binding assays. The intracellular location of GABARAP using chimeric GABARAP-GFP showed that the majority of GABARAP was located in perinuclear vesicles. Our results show that GABARAP plays a more general role outside the confines of neuronal cells and GABA(A) receptors.