Membrane-associated RING-CH (MARCH) 8 mediates the ubiquitination and lysosomal degradation of the transferrin receptor

The transferrin receptor (TfR) mediates the uptake of transferrin (Tf)-bound iron from the plasma into the cells of peripheral tissues. The TfR continuously recycles between the plasma membrane (PM) and early/recycling endosomes. TfR expression is tightly controlled by intracellular iron concentration through the regulation of TfR mRNA stability. However, much less is known about the mechanism by which TfR is degraded in cells. Previously, we reported a correlation between TfR ubiquitination and its iron-induced lysosomal degradation. The identification and characterization of a specific ubiquitin ligase for TfR is important to understand the mechanism of iron homeostasis. Here, we show that membrane-associated RING-CH (MARCH) 8 ubiquitinates TfR and promotes its lysosomal degradation. Similar to other RING-type ubiquitin ligases, the RING-CH domain of MARCH8, which is located in the N-terminal cytoplasmic (CT) domain, is essential for the ubiquitination and down-regulation of TfR. MARCH8 specifically recognizes the transmembrane (TM) domain of TfR and mediates ubiquitination of its CT domain. In addition, the six amino acid sequence located in the C-terminal CT domain of MARCH8, which is highly conserved among different species, is required for the down-regulation of TfR. Finally, and most importantly, TfR expression was markedly increased by siRNA-mediated knockdown of endogenous MARCH8. These findings demonstrate that the endogenous level of MARCH8 regulates TfR protein turnover via the down-regulation and ubiquitination of TfR.