Targeted locus amplification reveals heterogeneity between and within CFTR genotypes and association with CFTR function in patient-derived intestinal organoids.
CFTR
Haplotyping
Organoids
Phenotype-genotype relation
Journal
Journal of cystic fibrosis : official journal of the European Cystic Fibrosis Society
ISSN: 1873-5010
Titre abrégé: J Cyst Fibros
Pays: Netherlands
ID NLM: 101128966
Informations de publication
Date de publication:
May 2023
May 2023
Historique:
received:
23
12
2022
revised:
31
03
2023
accepted:
07
04
2023
medline:
12
6
2023
pubmed:
27
4
2023
entrez:
26
4
2023
Statut:
ppublish
Résumé
Cystic fibrosis (CF) disease severity can be highly variable, even between people with CF (pwCF) with similar genotypes. Here we use patient-derived intestinal organoids to study the influence of genetic variation within the cystic fibrosis transmembrane conductance regulator (CFTR) gene on CFTR function. Organoids of F508del/class I, F508del/S1251N and pwCF with only one detected CF-causing mutation were cultured. Allele-specific CFTR variation was investigated using targeted locus amplification (TLA), CFTR function was measured using the forskolin-induced swelling assay and mRNA levels were quantified using RT-qPCR. We were able to distinguish CFTR genotypes based on TLA data. Additionally, we observed heterogeneity within genotypes, which we were able to link to CFTR function for S1251N alleles. Our results indicate that the paired analysis of CFTR intragenic variation and CFTR function can gain insights in the underlying CFTR defect for individuals where the disease phenotype does not match the CFTR mutations detected during diagnosis.
Sections du résumé
BACKGROUND
BACKGROUND
Cystic fibrosis (CF) disease severity can be highly variable, even between people with CF (pwCF) with similar genotypes. Here we use patient-derived intestinal organoids to study the influence of genetic variation within the cystic fibrosis transmembrane conductance regulator (CFTR) gene on CFTR function.
METHODS
METHODS
Organoids of F508del/class I, F508del/S1251N and pwCF with only one detected CF-causing mutation were cultured. Allele-specific CFTR variation was investigated using targeted locus amplification (TLA), CFTR function was measured using the forskolin-induced swelling assay and mRNA levels were quantified using RT-qPCR.
RESULTS
RESULTS
We were able to distinguish CFTR genotypes based on TLA data. Additionally, we observed heterogeneity within genotypes, which we were able to link to CFTR function for S1251N alleles.
CONCLUSIONS
CONCLUSIONS
Our results indicate that the paired analysis of CFTR intragenic variation and CFTR function can gain insights in the underlying CFTR defect for individuals where the disease phenotype does not match the CFTR mutations detected during diagnosis.
Identifiants
pubmed: 37100706
pii: S1569-1993(23)00093-0
doi: 10.1016/j.jcf.2023.04.003
pii:
doi:
Substances chimiques
Cystic Fibrosis Transmembrane Conductance Regulator
126880-72-6
CFTR protein, human
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
538-547Informations de copyright
Copyright © 2023. Published by Elsevier B.V.