VACUOLAR H plus -ATPASE AS A NOVEL TARGET TO LOWER LOW DENSITY LIPOPROTEIN CHOLESTEROL

Objective: Low density lipoprotein cholesterol (LDL-c) is cleared from the circulation through binding to the hepatic LDL receptor (LDLR). The (pro)renin receptor [(P)RR, also known as vacuolar H+-ATPase (V-ATPase) accessory protein 2 or ATP6AP2)] is a regulator of this process. Indeed, (P)RR silencing markedly reduced LDLR protein abundance and thereby LDL uptake. Since V-ATPase is a multi-subunit complex (consisting of 13 subunits), in this study we evaluated the role of subunits other than the (P)RR. Design and method: siRNAs were designed against all V-ATPase subunits, transfected into HepG2 cells, and evaluated for their capacity to affect cell surface LDLR. Given that silencing subunit ATP6V1B2 upregulated LDLR most, we next designed an adeno-associated virus expressing shAtp6v1b2 and injected this into C57BL/6 mice and then investigated the effects of Atp6v1b2 knocking-down on LDL metabolism in vivo. Results: Silencing subunit ATP6V1B2 in HepG2 cells increased surface LDLR and total LDLR protein abundance by 1.5- and 2.3-fold, respectively. This was accompanied by both LDLR mRNA upregulation, suggestive for de novo protein synthesis, and enhanced LDL uptake following exposure to excess LDL. Silencing ATP6V1B2 significantly increased the mRNA levels of sterol regulatory element-binding protein 2 (SREBP2)-targeted genes (SQLE, HMGCR, and NPC1) as well as the nuclear SREBP2 protein level. In vivo, knocking-down Atp6v1b2 decreased plasma total cholesterol by 23%, without affecting total triglycerides. This was accompanied by LDLR upregulation in the liver, both at the mRNA and protein levels. Conclusions: The V-ATPase subunit ATP6V1B2 is an important regulator of LDLR expression. This involves the transcription factor SREBP2, and allows substantial LDL-c lowering. V-ATPase might be a novel pharmacological target to lower cholesterol.