Quantitative Proteomics of Human Fibroblasts with I1061T Mutation in Niemann–Pick C1 (NPC1) Protein Provides Insights into the Disease Pathogenesis*

Abstract

Niemann-Pick type C (NPC) disease is a fatal neurodegenerative disorder characterized by the accumulation of unesterified cholesterol in the late endosomal/lysosomal compartments. Mutations in the NPC1 protein are implicated in 95% of patients with NPC disease. The most prevalent mutation is the missense mutation I1061T that occurs in ϳ15-20% of the disease alleles. In our study, an isobaric labeling-based quantitative analysis of proteome of NPC1I1061T primary fibroblasts when compared with wild-type cells identified 281 differentially expressed proteins based on stringent data analysis criteria. Gene ontology enrichment analysis revealed that these proteins play important roles in diverse cellular processes such as protein maturation, energy metabolism, metabolism of reactive oxygen species, antioxidant activity, steroid metabolism, lipid localization, and apoptosis. The relative expression level of a subset of differentially expressed proteins (TOR4A, DHCR24, CLGN, SOD2, CHORDC1, HSPB7, and GAA) was independently and successfully substantiated by Western blotting. We observed that treating NPC1I1061T cells with four classes of seven different compounds that are potential NPC drugs increased the expression level of SOD2 and DHCR24. We have also shown an abnormal accumulation of glycogen in NPC1 I1061T fibroblasts possibly triggered by defective processing of lysosomal alpha-glucosidase. Our study provides a starting point for future more focused investigations to better understand the mechanisms by which the reported dysregulated proteins triggers the pathological cascade in NPC, and furthermore, their effect upon therapeutic interventions.

3.3 Å structure of Niemann–Pick C1 protein reveals insights into the function of the C-terminal luminal domain in cholesterol transport

“…The most frequent mutation, I1061T, present in 15-20% of all disease alleles (27,28), is in the α5 helix of the CTD (Fig. 4A).…”

Quantitative, Label‐Free Proteomics in the Symptomatic Niemann–Pick, Type C1 Mouse Model Using Standard Flow Liquid Chromatography and Thermal Focusing Electrospray Ionization

“…Other studies have reported secreted lysosomal enzyme alterations in both the mouse model and in NPC1 patients where many enzymes associated with drug metabolism have been identified that may have reduced activities in NPC1 patients . Cologna et al., identified potential markers of neuroinflammation in the cerebral cortex in both the mouse model as well as in NPC1 patients . Most recently, primary patient fibroblasts have been evaluated in large‐scale differential proteomics studies to reveal protein changes as a result of treatment with 2‐hydroxypropyl‐beta‐cyclodextrin or histone deacetylase inhibitors …”

Managing the Spatial Covariance of Genetic Diversity in Niemann-Pick C1 Through Modulation of the Hsp70 Chaperone System

“…In agreement with these data, it has also been shown that pharmacological treatments that lead to upregulation of NPC1 variant expression, including ryanodine receptor antagonists [17], treatment with oxysterols that bind to and stabilize NPC1 [18], reduced expression of TMEM97 [19], and HDAC inhibitors [20], partially abrogate the NPC1-I1061T phenotype in heterologous cell and patient fibroblast models. These findings highlight that modulation of the local folding and signaling environment can facilitate disease correction [21][22][23][24].…”