1. A trimeric Rab7 GEF controls NPC1-dependent lysosomal cholesterol export
- Author
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Ilana Berlin, Agata Sienkiewicz, Paul J. Lehner, Dick J. H. van den Boomen, Marlieke L.M. Jongsma, Daphne M. van Elsland, J. Paul Luzio, Jacques Neefjes, van den Boomen, Dick JH [0000-0001-6474-3661], Luzio, J Paul [0000-0003-3912-9760], Neefjes, Jacques JC [0000-0001-6763-2211], Lehner, Paul J [0000-0001-9383-1054], Apollo - University of Cambridge Repository, van den Boomen, Dick J. H. [0000-0001-6474-3661], Luzio, J. Paul [0000-0003-3912-9760], Neefjes, Jacques J. C. [0000-0001-6763-2211], and Lehner, Paul J. [0000-0001-9383-1054]
- Subjects
0301 basic medicine ,Hydroxymethylglutaryl-CoA Synthase ,49/47 ,General Physics and Astronomy ,Endogeny ,Membrane trafficking ,Small GTPases ,631/80/313/2011 ,chemistry.chemical_compound ,0302 clinical medicine ,Guanine Nucleotide Exchange Factors ,Homeostasis ,14/19 ,lcsh:Science ,Late endosome ,0303 health sciences ,Multidisciplinary ,631/80/313 ,030302 biochemistry & molecular biology ,Intracellular Signaling Peptides and Proteins ,3. Good health ,Cell biology ,13/31 ,Sterols ,Cholesterol ,631/45/287/1197 ,lipids (amino acids, peptides, and proteins) ,Protein Binding ,Endosome ,Science ,Protein subunit ,Endosomes ,Models, Biological ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,631/80/313/1624 ,Niemann-Pick C1 Protein ,Humans ,030304 developmental biology ,Fluorescent Dyes ,Genome, Human ,82/58 ,HEK 293 cells ,rab7 GTP-Binding Proteins ,Transporter ,Biological Transport ,General Chemistry ,Cholesterol, LDL ,030104 developmental biology ,HEK293 Cells ,chemistry ,Cholesterol import ,rab GTP-Binding Proteins ,Multiprotein Complexes ,LDL receptor ,lcsh:Q ,NPC1 ,CRISPR-Cas Systems ,Protein Multimerization ,Lysosomes ,030217 neurology & neurosurgery ,HeLa Cells - Abstract
Cholesterol import in mammalian cells is mediated by the LDL receptor pathway. Here, we perform a genome-wide CRISPR screen using an endogenous cholesterol reporter and identify >100 genes involved in LDL-cholesterol import. We characterise C18orf8 as a core subunit of the mammalian Mon1-Ccz1 guanidine exchange factor (GEF) for Rab7, required for complex stability and function. C18orf8-deficient cells lack Rab7 activation and show severe defects in late endosome morphology and endosomal LDL trafficking, resulting in cellular cholesterol deficiency. Unexpectedly, free cholesterol accumulates within swollen lysosomes, suggesting a critical defect in lysosomal cholesterol export. We find that active Rab7 interacts with the NPC1 cholesterol transporter and licenses lysosomal cholesterol export. This process is abolished in C18orf8-, Ccz1- and Mon1A/B-deficient cells and restored by a constitutively active Rab7. The trimeric Mon1-Ccz1-C18orf8 (MCC) GEF therefore plays a central role in cellular cholesterol homeostasis coordinating Rab7 activation, endosomal LDL trafficking and NPC1-dependent lysosomal cholesterol export., Lysosomes play an important role in cellular LDL-cholesterol uptake. Here, the authors perform a genome-wide genetic screen for cholesterol regulators and identify C18orf8 as a conserved subunit of a trimeric Rab7 GEF that controls LDL trafficking and NPC1-dependent lysosomal cholesterol export.
- Published
- 2019
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