@article{98ca6ec6e8ef4173898b824be271ae0d,
title = "NPC1 silent variant induces skipping of exon 11 (p.V562V) and unfolded protein response was found in a specific Niemann-Pick type C patient",
abstract = "BACKGROUND: Niemann-Pick type C (NPC, MIM \#257220) is a neuro-visceral disease, caused predominantly by pathogenic variants in the NPC1 gene. Here we studied patients with clinical diagnosis of NPC but inconclusive results regarding the molecular analysis.METHODS: We used a Next-Generation Sequencing (NGS)-panel followed by cDNA analysis. Latter, we used massively parallel single-cell RNA-seq (MARS-Seq) to address gene profiling changes and finally the effect of different variants on the protein and cellular levels.RESULTS: We identified novel variants and cDNA analysis allowed us to establish the functional effect of a silent variant, previously reported as a polymorphism. We demonstrated that this variant induces the skipping of exon 11 leading to a premature stop codon and identified it in NPC patients from two unrelated families. MARS-Seq analysis showed that a number of upregulated genes were related to the unfolded protein response (UPR) and endoplasmic reticulum (ER) stress in one specific patient. Also, for all analyzed variants, the NPC1 protein was partially retained in the ER.CONCLUSION: We showed that the NPC1 silent polymorphism (p.V562V) is a disease-causing variant in NPC and that the UPR is upregulated in an NPC patient.",
author = "Marisa Encarna{\c c}{\~a}o and Coutinho, \{Maria Francisca\} and Cho, \{Soo Min\} and Cardoso, \{Maria Teresa\} and Isaura Ribeiro and Paulo Chaves and Santos, \{Juliana In{\^e}s\} and Dulce Quelhas and L{\'u}cia Lacerda and \{Le{\~a}o Teles\}, Elisa and Futerman, \{Anthony H\} and Laura Vilarinho and Sandra Alves",
note = "Funding information: This work was partially supported by NORTE2020 (NORTE‐01‐0246‐FEDER‐000014 DESVENDAR “DEScobrir, VENcer as Doen{\c c}as Raras”, FCT (Funda{\c c}{\~a}o para a Ci{\^e}ncia e a Tecnologia ‐ MCTES, Portugal) projects PTDC/BBB‐BMD/6301/2014 and UIDB/00211/2020. MFC and JIS were grantees from the FCT (SFRH/BPD/101965/2014; SFRH/BD/124372/2016, respectively). We thank Dr. Paulo Matos from the confocal microscopy facility (INSA) for crucial help in the acquisition of confocal images, Dr. Paulo Gaspar (INSA) for discussion on biomarkers of the disease, Eug{\'e}nia Pinto (CGM), and Carla Caseiro (CGM) for technical help, the Bioinformatic department (WIS) for help in MARS‐seq analysis. Finally, we thank Prof Jos{\'e} Ramalho (CEDOC) and Dr. Olga Amaral (INSA) for kindly providing us with some antibodies. AUTHORS{\textquoteright} CONTRIBUTIONS MTC, PC, and ELT phenotyped the patient P1. ME and SA processed and analyzed the NGS‐data. ME, MFC, and IR performed the Sanger sequencing. ME performed RNA extractions, cDNA synthesis, qtRT‐PCR, WB, and IF. MFC and JIS performed the CHX treatments, some RNA extractions, and cell culture maintenance. IR performed filipin staining of all patients. SMC performed and analyzed RNA‐seq. ME, MFC, IR, SA, and SMC analyzed and interpreted the data. SA, LV, DQ, LL, and AHF obtained funding support. ME wrote the first version of the manuscript and prepared the figures. MFC helped in the first version of the manuscript. SA coordinated the work and corrected the manuscript. All authors read the final version of the manuscript and gave their permission for publication.",
year = "2020",
month = nov,
doi = "10.1002/mgg3.1451",
language = "الإنجليزيّة",
volume = "8",
pages = "e1451",
journal = "Molecular genetics \& genomic medicine",
issn = "2324-9269",
publisher = "John Wiley and Sons Inc.",
number = "11",
}