Posttranscriptional Regulation of the Human LDL Receptor by the U2-Spliceosome.
Paolo Zanoni, Grigorios Panteloglou, Alaa Othman, Joel T Haas, Roger Meier, Antoine Rimbert, Marta Futema, Yara Abou Khalil, Simon F Norrelykke, Andrzej J Rzepiela, Szymon Stoma, Michael Stebler, Freerk van Dijk, Melinde Wijers, Justina C Wolters, Nawar Dalila, Nicolette C A Huijkman, Marieke Smit, Antonio Gallo, Valérie Carreau, Anne Philippi, Jean-Pierre Rabès, Catherine Boileau, Michele Visentin, Luisa Vonghia, Jonas Weyler, Sven Francque, An Verrijken, Ann Verhaegen, Luc Van Gaal, Adriaan van der Graaf, Belle V van Rosmalen, Jerome Robert, Srividya Velagapudi, Mustafa Yalcinkaya, Michaela Keel, Silvija Radosavljevic, Andreas Geier, Anne Tybjaerg-Hansen, Mathilde Varret, Lucia Rohrer, Steve E Humphries, Bart Staels, Bart van de Sluis, Jan Albert Kuivenhoven, Arnold von Eckardstein
Journal: Circulation research
2022;130(1):80-95
PMID: 34809444
Abstract
BACKGROUND
The LDLR (low-density lipoprotein receptor) in the liver is the major determinant of LDL-cholesterol levels in human plasma. The discovery of genes that regulate the activity of LDLR helps to identify pathomechanisms of hypercholesterolemia and novel therapeutic targets against atherosclerotic cardiovascular disease.
METHODS
We performed a genome-wide RNA interference screen for genes limiting the uptake of fluorescent LDL into Huh-7 hepatocarcinoma cells. Top hit genes were validated by in vitro experiments as well as analyses of data sets on gene expression and variants in human populations.
RESULTS
The knockdown of 54 genes significantly inhibited LDL uptake. Fifteen of them encode for components or interactors of the U2-spliceosome. Knocking down any one of 11 out of 15 genes resulted in the selective retention of intron 3 of . The translated LDLR fragment lacks 88% of the full length LDLR and is detectable neither in nontransfected cells nor in human plasma. The hepatic expression of the intron 3 retention transcript is increased in nonalcoholic fatty liver disease as well as after bariatric surgery. Its expression in blood cells correlates with LDL-cholesterol and age. Single nucleotide polymorphisms and 3 rare variants of one spliceosome gene, , are associated with LDL-cholesterol in the population and familial hypercholesterolemia, respectively. Compared with overexpression of wild-type , overexpression of the 3 rare mutants in Huh-7 cells led to lower LDL uptake.
CONCLUSIONS
We identified a novel mechanism of posttranscriptional regulation of LDLR activity in humans and associations of genetic variants of with LDL-cholesterol levels.
Address:
Institute for Clinical Chemistry, University and University Hospital Zurich, Switzerland (P.Z., G.P., J.R., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Now with Institute of Medical Genetics, University of Zurich, Switzerland (P.Z.).; Center for Integrative Human Physiology, University of Zurich, Switzerland (P.Z., G.P., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Institute for Clinical Chemistry, University and University Hospital Zurich, Switzerland (P.Z., G.P., J.R., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Center for Integrative Human Physiology, University of Zurich, Switzerland (P.Z., G.P., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Institute of Molecular Systems Biology, ETH Zurich, Switzerland (A.O.).; University of Lille, Inserm, CHU Lille, Institut Pasteur de Lille, U1011- EGID, France (J.T.H., B.S.).; Scientific center for optical and electron microscopy (ScopeM), ETH Zurich, Switzerland (R.M., S.F.N., A.J.R., S.S., M. Stebler).; Department of Pediatrics, Section Molecular Genetics, University of Groningen, University Medical Center Groningen, the Netherlands (A.R., M.W., J.C.W., N.C.A.H., M. Smit, B.v.d.S., J.A.K.).; Now with Inserm UMR 1087/CNRS UMR 6291 IRS-UN, Nantes, France (A.R.).; Cardiology Research Centre, Molecular and Clinical Sciences Research Institute, St George's, University of London, United Kingdom (M.F.).; LVTS-INSERM UMRS 1148 and University of Paris, CHU Xavier Bichat, Paris, France (Y.A.K., J.-P.R., C.B., M. Varret).; Laboratory of Biochemistry and Molecular Therapeutics (LBTM), Faculty of Pharmacy and Pôle technologie Santé (PTS), Saint-Joseph University, Beirut, Lebanon (Y.A.K.).; Department of Genetics, University of Groningen, University Medical Center Groningen, the Netherlands (F.v.D., A.v.d.G.).; Department of Pediatrics, Section Molecular Genetics, University of Groningen, University Medical Center Groningen, the Netherlands (A.R., M.W., J.C.W., N.C.A.H., M. Smit, B.v.d.S., J.A.K.).; Department of Clinical Biochemistry, Rigshospitalet, Copenhagen University Hospital, Faculty of Health and Medical Sciences, University of Copenhagen, Denmark (N.D., A.T.-H.).; AP-HP, Endocrinology and Metabolism Department, Human Research Nutrition Center, Pitié-Salpêtrière Hospital, Paris, France (A. Gallo, V.C.).; Université de Paris, Faculté de Médecine Paris-Diderot, UMR-S958 Paris, France; Now with Université de Paris, Institut Cochin, INSERM U1016, CNRS UMR-8104, Paris, France (A.P.).; LVTS-INSERM UMRS 1148 and University of Paris, CHU Xavier Bichat, Paris, France (Y.A.K., J.-P.R., C.B., M. Varret).; AP-HP, Université Paris-Saclay, Paris, France (J.-P.R.).; UFR Simone Veil des Sciences de la Santé, UVSQ, Montigny-Le-Bretonneux, France (J.-P.R.).; LVTS-INSERM UMRS 1148 and University of Paris, CHU Xavier Bichat, Paris, France (Y.A.K., J.-P.R., C.B., M. Varret).; AP-HP, Genetics Department, CHU Xavier Bichat, Université de Paris, France (C.B.).; Department of Clinical Pharmacology and Toxicology, University Hospital Zurich, Switzerland (M. Visentin).; Department of Gastroenterology and Hepatology, Antwerp University Hospital, Edegem, Belgium (L.V., J.W., S.F.).; Laboratory of Experimental Medicine and Paediatrics, Faculty of Medicine, University of Antwerp, Belgium (L.V., J.W., S.F., A. Verrijken, A. Verhaegen, L.V.G.).; Department of Gastroenterology and Hepatology, Antwerp University Hospital, Edegem, Belgium (L.V., J.W., S.F.).; Laboratory of Experimental Medicine and Paediatrics, Faculty of Medicine, University of Antwerp, Belgium (L.V., J.W., S.F., A. Verrijken, A. Verhaegen, L.V.G.).; Department of Endocrinology, Diabetology and Metabolism, Antwerp University Hospital, Edegem, Belgium (A. Verrijken, A. Verhaegen, L.V.G.).; Department of Surgery, Academic Medical Center, University of Amsterdam, the Netherlands (B.V.v.R.).; Institute for Clinical Chemistry, University and University Hospital Zurich, Switzerland (P.Z., G.P., J.R., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Institute for Clinical Chemistry, University and University Hospital Zurich, Switzerland (P.Z., G.P., J.R., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Center for Molecular Cardiology, University of Zurich, Switzerland (S.V.).; Center for Integrative Human Physiology, University of Zurich, Switzerland (P.Z., G.P., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Institute for Clinical Chemistry, University and University Hospital Zurich, Switzerland (P.Z., G.P., J.R., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Division of Molecular Medicine, Department of Medicine, Columbia University, New York, NY (M.Y.).; Center for Integrative Human Physiology, University of Zurich, Switzerland (P.Z., G.P., S.V., M.Y., M.K., S.R., L.R., A.v.E.).; Division of Hepatology, Department of Medicine II, University Hospital Würzburg, Germany (A. Geier).; LVTS-INSERM UMRS 1148 and University of Paris, CHU Xavier Bichat, Paris, France (Y.A.K., J.-P.R., C.B., M. Varret).; Cardiovascular Genetics, Institute of Cardiovascular Science, University College London, United Kingdom (S.E.H.).
Link outs
Free resources
Full Text Sources:
Molecular Biology Databases:
Subscription / membership required
MeSH Terms:
Cholesterol,
HEK293 Cells,
Hep G2 Cells,
Humans,
Lipoproteins, LDL,
Liver,
Mutation,
Nuclear Proteins,
RNA Splicing,
Receptors, LDL,
Spliceosomes
