β-Estradiol results in a proprotein convertase subtilisin/kexin type 9-dependent increase in low-density lipoprotein receptor levels in human hepatic HuH7 cells.
The lower risk of coronary artery disease in premenopausal women than in men and postmenopausal women implicates sex steroids in cardioprotective processes. β-Estradiol upregulates liver low-density lipoprotein receptor (LDLR), which, in turn, decreases circulating levels of low-density lipoprotein, which is a risk factor for coronary artery disease. Conversely, LDLR protein is negatively regulated by proprotein convertase subtilisin/kexin type 9 (PCSK9). Herein, we investigated PCSK9 regulation by β-estradiol and its impact on LDLR in human hepatocarcinoma HuH7 cells grown in the presence or absence of β-estradiol. Immunoblot analysis showed upregulation of LDLR at 3 μm β-estradiol (140%), and the upregulation reached 220% at 10 μm β-estradiol; only at the latter dose was an increase in LDLR mRNA detected by qPCR, suggesting post-translational regulation of LDLR. No changes in PCSK9 mRNA or secreted protein levels were detected by qPCR or ELISA, respectively. β-estradiol-conditioned medium devoid of PCSK9 failed to upregulate LDLR. Similarly, PCSK9 knockdown cells showed no upregulation of LDLR by β-estradiol. Together, these results indicate a requirement for PCSK9 in the β-estradiol-induced upregulation of LDLR. A radiolabeling assay showed a significant, dose-dependent decrease in the ratio of secreted phosphoPCSK9 to total secreted PCSK9 with increasing β-estradiol levels, suggesting a change in the functional state of PCSK9 in the presence of β-estradiol. Our results indicate that the protein upregulation of LDLR at subtranscriptionally effective doses of β-estradiol, and its supratranscriptional upregulation at 10 μm β-estradiol, occur through an extracellular PCSK9-dependent mechanism.
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Amanda E
AE
Starr
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Valérie
V
Lemieux
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Jenny
J
Noad
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Jasmine I
JI
Moore
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Thilina
T
Dewpura
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Angela
A
Raymond
Chronic Disease Program, Ottawa Hospital Research Institute, The Ottawa Hospital, Ontario, Canada.|Laboratory of Biochemical Neuroendocrinology, Clinical Research Institute of Montreal, Quebec, Canada.
Michel
M
Chrétien
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Daniel
D
Figeys
Ottawa Institute of Systems Biology, Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Janice
J
Mayne
D002460:Cell Line / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004958:Estradiol / Q000378:metabolism* / Q000494:pharmacology; D056945:Hep G2 Cells / Q000187:drug effects; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases / Q000235:genetics / Q000378:metabolism*; D011973:Receptors, LDL / Q000235:genetics / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
D011973:Receptors, LDL; D004958:Estradiol; C472125:PCSK9 protein, human; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases; D012697:Serine Endopeptidases
atherosclerosis; estrogen; lipoproteins/receptors; low-density lipoprotein receptor; proprotein convertase subtilisin/kexin type 9