The transferrin receptor-1 membrane stub undergoes intramembrane proteolysis by signal peptide peptidase-like 2b

Abstract

The successive events of shedding and regulated intramembrane proteolysis are known to comprise a fundamental biological process of type I and II membrane proteins (e.g. amyloid precursor protein, Notch receptor and pro-tumor necrosis factor-α). Some of the resulting fragments were shown to be involved in important intra- and extracellular signalling events. Although shedding of the human transferrin receptor-1 (TfR1) has been known for > 30 years and soluble TfR1 is an accepted diagnostic marker, the fate of the remaining N-terminal fragment (NTF) remains unknown. In the present study, we demonstrate for the first time that TfR1-NTF is subject to regulated intramembrane proteolysis and, using MALDI-TOF-TOF-MS, we have identified the cleavage site as being located C-terminal from Gly-84. We showed that the resulting C-terminal peptide is extracellularly released after regulated intramembrane proteolysis and it was detected as a monomer with an internal disulfide bridge. We further identified signal peptide peptidase-like 2a and mainly signal peptide peptidase-like 2b as being responsible for the intramembrane proteolysis of TfR1-NTF. Shedding and regulated intramembrane proteolysis (RIP) are fundamental biological processes of membrane proteins. The resulting fragments are involved in important intra- and extracellular signaling events. We demonstrate that the transferrin receptor is subject to RIP and identified the protease SPPL2b being responsible for cleavage at Gly-84. The resulting C-peptide is extracellularly released as a monomer with an internal disulfide bridge. © 2013 FEBS.