APOB CRISPR-Cas9 Engineering in Hypobetalipoproteinemia: A Promising Tool for Functional Studies of Novel Variants
2022
Vanhoye, Xavier | Janin, Alexandre | Caillaud, Amandine | Rimbert, Antoine | Venet, Fabienne | Gossez, Morgane | Dijk, Wieneke | Marmontel, Oriane | Nony, Séverine | Chatelain, Charlotte | Durand, Christine | Lindenbaum, Pierre | Rieusset, J. | Cariou, Bertrand | Moulin, Philippe | Di Filippo, Mathilde | Hospices Civils de Lyon (HCL) | Institut NeuroMyoGène (INMG) ; Université Claude Bernard Lyon 1 (UCBL) ; Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS) | ITX-lab unité de recherche de l'institut du thorax UMR1087 UMR6291 (ITX-lab) ; Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Nantes Université - UFR de Médecine et des Techniques Médicales (Nantes Univ - UFR MEDECINE) ; Nantes Université - pôle Santé ; Nantes Université (Nantes Univ)-Nantes Université (Nantes Univ)-Nantes Université - pôle Santé ; Nantes Université (Nantes Univ)-Nantes Université (Nantes Univ) | Hôpital Edouard Herriot [CHU - HCL] ; Hospices Civils de Lyon (HCL) | Centre International de Recherche en Infectiologie (CIRI) ; École normale supérieure de Lyon (ENS de Lyon) ; Université de Lyon-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL) ; Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS) | Cardiovasculaire, métabolisme, diabétologie et nutrition (CarMeN) ; Université Claude Bernard Lyon 1 (UCBL) ; Université de Lyon-Université de Lyon-Hospices Civils de Lyon (HCL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE) | Hôpital Louis Pradel [CHU - HCL] ; Hospices Civils de Lyon (HCL) | ANR-16-RHUS-0007,CHOPIN,CHOPIN(2016)
International audience
اظهر المزيد [+] اقل [-]إنجليزي. Hypobetalipoproteinemia is characterized by LDL-cholesterol and apolipoprotein B (apoB) plasma levels below the fifth percentile for age and sex. Familial hypobetalipoproteinemia (FHBL) is mostly caused by premature termination codons in the APOB gene, a condition associated with fatty liver and steatohepatitis. Nevertheless, many families with a FHBL phenotype carry APOB missense variants of uncertain significance (VUS). We here aimed to develop a proof-of-principle experiment to assess the pathogenicity of VUS using the genome editing of human liver cells. We identified a novel heterozygous APOB-VUS (p.Leu351Arg), in a FHBL family. We generated APOB knock-out (KO) and APOB-p.Leu351Arg knock-in Huh7 cells using CRISPR-Cas9 technology and studied the APOB expression, synthesis and secretion by digital droplet PCR and ELISA quantification. The APOB expression was decreased by 70% in the heterozygous APOB-KO cells and almost abolished in the homozygous-KO cells, with a consistent decrease in apoB production and secretion. The APOB-p.Leu351Arg homozygous cells presented with a 40% decreased APOB expression and undetectable apoB levels in cellular extracts and supernatant. Thus, the p.Leu351Arg affected the apoB secretion, which led us to classify this new variant as likely pathogenic and to set up a hepatic follow-up in this family. Therefore, the functional assessment of APOB-missense variants, using gene-editing technologies, will lead to improvements in the molecular diagnosis of FHBL and the personalized follow-up of these patients.
اظهر المزيد [+] اقل [-]الكلمات المفتاحية الخاصة بالمكنز الزراعي (أجروفوك)
المعلومات البيبليوغرافية
تم تزويد هذا السجل من قبل Institut national de la recherche agronomique