A mouse model for the cystic fibrosis delta F508 mutation.

Abstract

Most cystic fibrosis (CF) patients produce a mutant form (delta F508) of the cystic fibrosis transmembrane conductance regulator (CFTR), which is not properly processed in normal cells but is active as a chloride channel in several experimental systems. We used a double homologous recombination (‘Hit and Run’) procedure to generate a mouse model for the delta F508 mutation. Targeted embryonic stem (ES) cells (Hit clones) were found; of these either 80 or 20% of the clones had lost the delta F508 mutation, depending on the distance between the linearization site in the targeting construct and the delta F508 mutation. Correctly targeted clones underwent a second selection step resulting in ES cell clones (Run clones) heterozygous for the delta F508 mutation with an efficiency of 2–7%. Chimeric mice were generated and offspring homozygous for the delta F508 mutation showed electrophysiological abnormalities in nasal epithelium, gallbladder and in the intestine, and histological abnormalities in the intestine, typical of CF. Our data suggest that the delta F508 mice have residual delta F508 CFTR activity which would explain the mild pathology of the delta F508 mice. The delta F508 mouse may provide a useful model for the study of the processing defect of delta F508 CFTR and for the development of novel therapeutic approaches based on circumvention of the processing block.

Mutations in the Nucleotide Binding Domain 1 Signature Motif Region Rescue Processing and Functional Defects of Cystic Fibrosis Transmembrane Conductance Regulator ΔF508

“…7). A significant level of CFTR⌬F508 chloride channel activity and a detectable level of mature protein have been observed for the homozygous ⌬F508 CF mouse (65)(66)(67)(68). Our results suggest that the T539 could contribute to the attenuated defect caused by the ⌬F508 mutation in the murine CFTR.…”

Cystic Fibrosis Is Associated with a Defect in Apical Receptor–Mediated Endocytosis in Mouse and Human Kidney

“…However, ⌬F508-CFTR can function as a cAMP-regulated chloride channel, both in the plasma membrane and intracellularly (27). Previous studies demonstrated a residual chloride permeability in intestine and gallbladder of the Cftr ⌬F/⌬F mice used here (24), and the electrolyte and water handling is preserved in the PT of Cftr tm2cam ⌬F508…”

“…These data suggest that in Cftr Ϫ/Ϫ mice, the complete loss of CFTR induces a significant decrease in the membrane stability and increased urinary excretion of cubilin, leading to a defective apical receptor-mediated endocytosis in PT cells, with urinary loss of LMW ligands (␤ 2 -microglobulin, CC16, and transferrin). This phenotype is observed less consistently in Cftr ⌬F/⌬F mice, which may be explained by a variable residual expression and/or functionality of the mutated ⌬F508-CFTR in the Cftr ⌬F/⌬F kidneys (24,33).…”

Targeting autophagy as a novel strategy for facilitating the therapeutic action of potentiators on ΔF508 cystic fibrosis transmembrane conductance regulator

“…34 Therefore, we aimed at translating these findings in vivo, into a mouse model of CF. For this, we took advantage of mice homozygous for the ΔF508-CFTR mutation in the 129/FVB outbred background (Cftrtm1EUR, F508del, FVB/129) (Cftr F508del ), an established animal model of CF, [46][47][48] that shows constitutive lung inflammation. 7,48 First, Cftr F508del mice were transduced intranasally with pLKO.1-CMV-tGFP lentiviral vectors expressing mouse shRNA specific for Sqstm1 or no target sequence (control shRNA) (n = 5 for each group of treatment) ( Fig.…”