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Transcript, protein, metabolite and cellular studies in skin fibroblasts demonstrate variable pathogenic impacts of NPC1 mutations.

ABSTRACT: BACKGROUND:Niemann-Pick type C (NP-C) is a rare neurovisceral genetic disorder caused by mutations in the NPC1 or the NPC2 gene. NPC1 is a multipass-transmembrane protein essential for egress of cholesterol from late endosomes/lysosomes. To evaluate impacts of NPC1 mutations, we examined fibroblast cultures from 26 NP-C1 patients with clinical phenotypes ranging from infantile to adult neurologic onset forms. The cells were tested with multiple assays including NPC1 mRNA expression levels and allele expression ratios, assessment of NPC1 promoter haplotypes, NPC1 protein levels, cellular cholesterol staining, localization of the mutant NPC1 proteins to lysosomes, and cholesterol/cholesteryl ester ratios. These results were correlated with phenotypes of the individual patients. RESULTS:Overall we identified 5 variant promoter haplotypes. Three of them showed reporter activity decreased down to 70% of the control sequence. None of the haplotypes were consistently associated with more severe clinical presentation of NP-C. Levels of transcripts carrying null NPC1 alleles were profoundly lower than levels of the missense variants. Low levels of the mutant NPC1 protein were identified in most samples. The protein localised to lysosomes in cultures expressing medium to normal NPC1 levels. Fibroblasts from patients with severe infantile phenotypes had higher cholesterol levels and higher cholesterol/cholesteryl ester ratios. On the contrary, cell lines from patients with juvenile and adolescent/adult phenotypes showed values comparable to controls. CONCLUSION:No single assay fully correlated with the disease severity. However, low residual levels of NPC1 protein and high cholesterol/cholesteryl ester ratios associated with severe disease. The results suggest not only low NPC1 expression due to non-sense mediated decay or low mutant protein stability, but also dysfunction of the stable mutant NPC1 as contributors to the intracellular lipid transport defect.

SUBMITTER: Musalkova D

PROVIDER: S-EPMC7132889 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

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Transcript, protein, metabolite and cellular studies in skin fibroblasts demonstrate variable pathogenic impacts of NPC1 mutations.

Musalkova Dita D Majer Filip F Kuchar Ladislav L Luksan Ondrej O Asfaw Befekadu B Vlaskova Hana H Storkanova Gabriela G Reboun Martin M Poupetova Helena H Jahnova Helena H Hulkova Helena H Ledvinova Jana J Dvorakova Lenka L Sikora Jakub J Jirsa Milan M Vanier Marie T MT Hrebicek Martin M

Orphanet journal of rare diseases 20200405 1

<h4>Background</h4>Niemann-Pick type C (NP-C) is a rare neurovisceral genetic disorder caused by mutations in the NPC1 or the NPC2 gene. NPC1 is a multipass-transmembrane protein essential for egress of cholesterol from late endosomes/lysosomes. To evaluate impacts of NPC1 mutations, we examined fibroblast cultures from 26 NP-C1 patients with clinical phenotypes ranging from infantile to adult neurologic onset forms. The cells were tested with multiple assays including NPC1 mRNA expression level ...[more]

PMID: 32248828

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Dataset Information

Transcript, protein, metabolite and cellular studies in skin fibroblasts demonstrate variable pathogenic impacts of NPC1 mutations.

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Transcript, protein, metabolite and cellular studies in skin fibroblasts demonstrate variable pathogenic impacts of NPC1 mutations.

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