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bioRxiv · 10.1101/835686

A trimeric Rab7 GEF controls NPC1-dependent lysosomal cholesterol export

van den Boomen, D. J.·Sienkiewicz, A.·Berlin, I.·Jongsma, M. L.·van Elsland, D. M.·Luzio, J. P.·Neefjes, J. J.·Lehner, P. J.
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Abstract

Cholesterol import in mammalian cells is mediated by the LDL receptor pathway. Here, using an endogenous cholesterol reporter in a genome-wide CRISPR screen we identify >70 genes involved in LDL-cholesterol import. We characterise C18orf8 as a core component 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 additional 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.

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van den Boomen, D. J., Sienkiewicz, A., Berlin, I., Jongsma, M. L., van Elsland, D. M., Luzio, J. P., Neefjes, J. J., Lehner, P. J.. 2019-11-08. A trimeric Rab7 GEF controls NPC1-dependent lysosomal cholesterol export. https://doi.org/10.1101/835686

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