Abstract

NPC1 gene encodes a transmembrane glycoprotein on the late endosome/lysosomal membrane. Its mutation leads to a rare and aggravated autosomal recessive neurovisceral condition, termed Niemann-Pick disease type C1 (NPC1), which is characterized by progressive neurodegeneration, visceral symptoms, and premature death. To investigate the influence of NPC1 gene deletion on cell morphology, adhesion, proliferation, and apoptosis, CRISPR-Cas9 technology was used to knockout the NPC1 gene in HEK 293 T cells. Sanger sequencing, western blotting, and immunofluorescence were used to confirm successful NPC1 ablation. Filipin staining results indicated that deletion of NPC1 gene led to accumulation of unesterified cholesterol in HEK 293 T cells. Phalloidin staining results revealed cell aggregation, synapse shortening, nuclear enlargement, and cytoskeleton filamentous actin thinning in HEK 293 T cells with NPC1 gene mutation. Furthermore, NPC1 gene mutated HEK 293 T cell showed enhanced cell adhesion, inhibited cell proliferation, and increased cell apoptosis. In addition, NPC1 gene mutations significantly increased the protein expression levels of E-cadherin and γ-catenin and significantly decreased the protein expression levels of Wnt 3a, c-Myc, and cyclin D1. These results suggest that NPC1 may regulate cell adhesion by affecting the cadherin–catenin complex through E-cadherin, and that the classical Wnt signaling pathway may be inhibited by restricting β-catenin from entering the nucleus to inhibit cell proliferation.