Hepatic PCSK9 Expression Is Regulated by Nutritional Status via Insulin and Sterol Regulatory Element-binding Protein 1c

Philippe Costet,Bertrand Cariou,Gilles Lambert,Florent Lalanne,Bernard Lardeux,Anne-Laure Jarnoux,Aldo Grefhorst,Bart Staels,Michel Krempf

doi:10.1074/jbc.m508582200

Abstract

Familial autosomal dominant hypercholesterolemia is associated with high risk for cardiovascular accidents and is related to mutations in the low density lipoprotein receptor or its ligand apolipoprotein B (apoB). Mutations in a third gene, proprotein convertase subtilisin kexin 9 (PCSK9), were recently associated to this disease. PCSK9 acts as a natural inhibitor of the low density lipoprotein receptor pathway, and both genes are regulated by depletion of cholesterol cell content and statins, via sterol regulatory element-binding protein (SREBP). Here we investigated the regulation of PCSK9 gene expression during nutritional changes. We showed that PCSK9 mRNA quantity is decreased by 73% in mice after 24 h of fasting, leading to a 2-fold decrease in protein level. In contrast PCSK9 expression was restored upon high carbohydrate refeeding. PCSK9 mRNA increased by 4-5-fold in presence of insulin in rodent primary hepatocytes, whereas glucose had no effect. Moreover, insulin up-regulated hepatic PCSK9 expression in vivo during a hyperinsulinemic-euglycemic clamp in mice. Adenoviral mediated overexpression of a dominant or negative form of SREBP-1c confirmed the implication of this transcription factor in insulin-mediated stimulation of PCSK9 expression. Liver X receptor agonist T0901317 also regulated PCSK9 expression via this same pathway (a 2-fold increase in PCSK9 mRNA of primary hepatocytes cultured for 24 h in presence of 1 microm T0901317). As our last investigation, we isolated PCSK9 proximal promoter and verified the functionality of a SREBP-1c responsive element located from 335 bp to 355 bp upstream of the ATG. Together, these results show that PCSK9 expression is regulated by nutritional status and insulinemia.

Highlights

Another form of this disease, familial defective apolipoproteinB100, is caused by mutations in the LDL receptor (LDLr) binding domain of ApoB100 [1]
Hepatic proprotein convertase subtilisin kexin 9 (PCSK9) Expression Is Decreased by Fasting and Restored upon Refeeding—To determine whether PCSK9 expression is modulated by glycemia and/or insulinemia, C57BL/6 female mice were subjected to a 24-h fasting period followed by refeeding with a carbohydrate-rich diet for 6 and 24 h; their glycemia and insulinemia were determined at each time point, and PCSK9 mRNA and protein levels were analyzed afterward (Fig. 1)
PCSK9 expression was restored by refeeding, showing appreciable levels (27% of control fed mice) as soon as 6 h after refeeding; mRNA levels subsequently returned to their non-fasted state after 24 h of refeeding

Summary

Introduction

Another form of this disease, familial defective apolipoprotein (apo)B100, is caused by mutations in the LDLr binding domain of ApoB100 [1]. The transcription of both genes is activated by cholesterol cell content depletion, via the sterol regulatory element-binding protein (SREBP) [11,12,13,14]. PCSK9 is down-regulated by dietary cholesterol, and moderately up-regulated by the liver X receptor (LXR) agonist T0901317. When cellular sterols are low, SREBP-1a and SREBP-2 are cleaved into a mature, truncated, peptide that is directed to the nucleus, where it activates its target genes. LXR is involved in many steps of reverse cholesterol transport, macrophage cholesterol efflux, fecal cholesterol excretion, and bile acid synthesis. It constitutes a promising therapeutic target [31]. These data, together with the ability of T0901317 to activate PCSK9 in vivo, which could be an indirect effect mediated by SREBP-1c, prompted us to investigate whether insulin could activate PCSK9 transcription via SREBP-1c

Methods

Results

Conclusion