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J Biol Chem. 2018 Oct 5;293(40):15556-15568. doi: 10.1074/jbc.RA118.002640. Epub 2018 Aug 24.

Membrane cholesterol as regulator of human rhomboid protease RHBDL4.

Author information

1
From the Department of Pharmacology and Therapeutics and Cell Information Systems Group and.
2
Department of Biochemistry and Groupe de Recherche Axé sur la Structure des Protéines, McGill University, Montreal, Quebec H3G 0B1, Canada and.
3
the Department of Physics, Theoretical Molecular Biophysics, Freie Universität Berlin, Arnimallee 14, Berlin 14195, Germany.
4
From the Department of Pharmacology and Therapeutics and Cell Information Systems Group and lisa.munter@mcgill.ca.

Abstract

In the last decade, intramembrane proteases have gained increasing attention because of their many links to various diseases. Nevertheless, our understanding as to how they function or how they are regulated is still limited, especially when it comes to human homologues. In this regard, here we sought to unravel mechanisms of regulation of the protease rhomboid-like protein-4 (RHBDL4), one of five active human serine intramembrane proteases. In view of our recent finding that human RHBDL4 efficiently cleaves the amyloid precursor protein (APP), a key protein in the pathology of Alzheimer's disease, we used established reagents to modulate the cellular cholesterol content and analyzed the effects of this modulation on RHBDL4-mediated processing of endogenous APP. We discovered that lowering membrane cholesterol levels increased the levels of RHBDL4-specific endogenous APP fragments, whereas high cholesterol levels had the opposite effect. Direct binding of cholesterol to APP did not mediate these modulating effects of cholesterol. Instead, using homology modeling, we identified two potential cholesterol-binding motifs in the transmembrane helices 3 and 6 of RHBDL4. Substitution of the essential tyrosine residues of the potential cholesterol-binding motifs to alanine increased the levels of endogenous APP C-terminal fragments, reflecting enhanced RHBDL4 activity. In summary, we provide evidence that the activity of RHBDL4 is regulated by cholesterol likely through a direct binding of cholesterol to the enzyme.

KEYWORDS:

Alzheimer's disease; amyloid precursor protein (APP); cholesterol; cholesterol pulldown; cholesterol recognition amino acid consensus sequence (CRAC); cholesterol-binding protein; intramembrane proteolysis; large APP C-terminal fragments; low-density lipoprotein (LDL); neurodegeneration; rhomboid protease

PMID:
30143535
PMCID:
PMC6177575
[Available on 2019-10-05]
DOI:
10.1074/jbc.RA118.002640
[Indexed for MEDLINE]

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