Abstract
Cholesterol accumulation in late endosomes is a prevailing phenotype of Niemann-Pick type C1 (NPC1) mutant cells. Likewise, annexin A6 (AnxA6) overexpression induces a phenotype reminiscent of NPC1 mutant cells. Here, we demonstrate that this cellular cholesterol imbalance is due to AnxA6 promoting Rab7 inactivation via TBC1D15, a Rab7-GAP. In NPC1 mutant cells, AnxA6 depletion and eventual Rab7 activation was associated with peripheral distribution and increased mobility of late endosomes. This was accompanied by an enhanced lipid accumulation in lipid droplets in an acyl-CoA:cholesterol acyltransferase (ACAT)-dependent manner. Moreover, in AnxA6-deficient NPC1 mutant cells, Rab7-mediated rescue of late endosome-cholesterol export required the StAR-related lipid transfer domain-3 (StARD3) protein. Electron microscopy revealed a significant increase of membrane contact sites (MCS) between late endosomes and ER in NPC1 mutant cells lacking AnxA6, suggesting late endosome-cholesterol transfer to the ER via Rab7 and StARD3-dependent MCS formation. This study identifies AnxA6 as a novel gatekeeper that controls cellular distribution of late endosome-cholesterol via regulation of a Rab7-GAP and MCS formation.
Highlights
The transmembrane Niemann-Pick type C1 (NPC1) protein is essential for the efficient export of cholesterol from endolysosomes
We previously demonstrated that annexin A6 (AnxA6) overexpression led to late endosome-cholesterol accumulation, a phenotype reminiscent of the NPC1 mutant phenotype [46, 62]
This was accompanied by an increased recruitment of AnxA6 to cholesterol-laden late endosomes upon pharmacological NPC1 inhibition, using U18666A, or loading with low-density lipoproteins (LDL) [43,44,45]
Summary
The transmembrane NPC1 protein is essential for the efficient export of cholesterol from endolysosomes. Several other cytoplasmic players contribute, via vesicular and/or non-vesicular pathways, to the exit of cholesterol from this compartment [1,2,3], including members of the oxysterol-binding protein (OSBP) family, such as oxysterol-related protein 1L (ORP1L), the small GTPases Rab, Rab and Rab, as well as the late endosome, membrane-anchored StARD3 and StARD3 N-terminal like (StARD3NL) proteins [3,4,5,6]. Some of the cytoplasmic proteins that bind and transport cholesterol are engaged in the formation and functions of membrane contact sites (MCS) that are emerging as important non-vesicular transfer mediators for lipids, cholesterol or calcium ( Ca2+) between compartments [7,8,9,10,11,12,13,14,15,16,17,18,19,20]. Despite its participation in cholesterol transfer between compartments via MCS, StARD3 overexpression did not increase cholesterol esterification via acyl-CoA:cholesterol acyltransferase (ACAT) in the ER [31, 32] and was unable to rescue late endosome-cholesterol accumulation in NPC1 mutant cells [23, 24]
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