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 Flyvbjerg Nørrelykke, Andrzej Rzepiela, Szymon Stoma, Michael Stebler, Freerk van Dijk, Melinde Wijers, Justina C Wolters, Nawar Dalila, Nicolette Huijkman, Marieke Smit, Antonio Gallo, Valerie CarreauAnne Philippi, Jean-Pierre Rabès, Catherine Boileau, Michele Visentin, Luisa Vonghia, Jonas Weyler, Sven Francque, An Verrijken, Ann Verhaegen, Luc F Van Gaal, Adriaan van der Graaf, Belle V van Rosmalen, Jérôme Robert, Srividya Velagapudi, Mustafa Yalcinkaya, Michaela Keel, Silvija Radosavljevic, Andreas Geier, Anne Tybjærg-Hansen, Mathilde Varret, Lucia Rohrer, Steve E Humphries, Bart Staels, Bart van de Sluis, Jan Albert Kuivenhoven, Arnold Von Eckardstein*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

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Abstract

Background: The low-density lipoprotein receptor (LDLR) 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 datasets 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 LDLR. The translated LDLR fragment lacks 88% of the full length LDLR and is detectable neither in non-transfected cells nor in human plasma. The hepatic expression of the intron 3 retention transcript is increased in non-alcoholic fatty liver disease as well as after bariatric surgery. Its expression in blood cells correlates with LDL-cholesterol and age. Single nucleotide polymorphisms and three rare variants of one spliceosome gene, RBM25, are associated with LDL-cholesterol in the population and familial hypercholesterolemia, respectively. Compared to overexpression of wild type RBM25, overexpression of the three rare RBM25 mutants in Huh-7 cells led to lower LDL uptake.

Conclusions: We identified a novel mechanism of post-transcriptional regulation of LDLR activity in humans and associations of genetic variants of RBM25 with LDL-cholesterol levels.

Original languageEnglish
Pages (from-to)80-95
JournalCirculation research
Volume130
Issue number1
Early online date23-Nov-2021
DOIs
Publication statusPublished - 7-Jan-2022

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