The multidrug resistance pump ABCB1 is a substrate for the ubiquitin ligase NEDD4-1. (March 2015)

• Record Type: Journal Article
• Authors: Akkaya, Begum G.; Zolnerciks, Joseph K.; Ritchie, Tasha K.; Bauer, Bjoern; Hartz, Anika M. S.; Sullivan, James A.; Linton, Kenneth J.
• Is Part Of: Molecular membrane biology. Volume 32:Number 2(2015)
• Page Start: 39
• Page End: 45
• Publication Date: 2015-03
• Subjects: Membranes (Biology) -- Periodicals; Biochemistry -- Periodicals; Cell membranes -- Periodicals; Molecular biology -- Periodicals; 571.64
• Languages: English
• ISSNs: 0968-7688

Abstract

<abstract> <title>Abstract</title> <p>The ATP Binding Cassette transporter ABCB1 can export the neurotoxic peptide β-amyloid from endothelial cells that line the blood-brain barrier (BBB). This has the potential to lower cerebral levels of β-amyloid, but ABCB1 expression in the BBB appears to be progressively reduced in patients with Alzheimer's disease. The surface density of many membrane proteins is regulated by ubiquitination catalyzed by ubiquitin E3 ligases. In brain capillaries of mice challenged with β-amyloid <italic>ex vivo</italic>, we show that the level of the ubiquitin ligase Nedd4 increases concomitant with reduction in Abcb1. <italic>In vitro</italic> we show that human ABCB1 is a substrate for human NEDD4-1 ligase. Recombinant ABCB1 was purified from Sf21 insect cells and incubated with recombinant NEDD4-1 purified from <italic>Escherichia coli</italic>. The treated ABCB1 had reduced mobility on SDS-PAGE, and mass spectrometry identified eight lysine residues, K271, K272, K575, K685, K877, K885, K887 and K1062 that were ubiquitinated by NEDD4-1. Molecular modelling showed that all of the residues are exposed on the surface of the intracellular domains of ABCB1. K877, K885 and K887 in particular, are located in the intracellular loop of transmembrane helix 10 (TMH10) in close proximity, in the tertiary fold, to a putative NEDD4-1 binding site in the intracellular helix extending from TMH12 (PxY motif, residues 996–998). Transient expression of NEDD4-1 in HEK293 Flp-In cells stably expressing ABCB1 was shown to reduce the surface density of the transporter. Together, the data identify this ubiquitin ligase as a potential target for intervention in the pathophysiology of Alzheimer's disease.</p> </abstract>