The Function of C19orf12 and the Cytotoxic Mechanism of Its Pathogenic Mutants
Objective:This study aims to investigate the role of the neurodegenerative disease gene C19orf12 in the H1299 cell line and to analyze the potential impact of its pathogenic mutant(p.Ala63Pro).Methods:The expression levels of C19orf12 in various cell lines,including Hela,A549,U2OS,and H1299,were analyzed using Reverse Transcription Quantitative PCR(RT-qPCR)and West-ern Blot.An shRNA lentiviral expression vector targeting C19orf12 was constructed,and C19orf12 was knocked down in the H1299 cell line.Data-independent acquisition(DIA)proteomics was utilized to analyze the effect of C19orf12 knockdown on the cellular protein ex-pression profile.Differentially expressed proteins were subjected to protein-protein interaction(PPI)network analysis,and the MCODE tool was used to identify key protein interaction sub-networks.Finally,immunofluorescence and co-immunoprecipitation were performed to analyze the subcellular localization and interaction changes of the C19orf12 mutant proteins.Results:C19orf12 was expressed at rela-tively high levels in H1299 cells and its knockdown significantly inhibited cell growth.Proteomic analysis identified 216 significantly differentially expressed proteins,with 132 upregulated and 84 downregulated.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analyses revealed that these differentially expressed proteins were mainly involved in pathways related to neurodegenerative diseases such as Parkinson's disease,Alzheimer's disease,and Huntington's disease,as well as mitochondrial oxidative phosphorylation,lipid metabolism,and heart disease-related pathways.The pathogenic mutant of C19orf12(p.Ala63Pro)lost the ability to interact with multiple key proteins.Conclusion:C19orf12 plays a key role in cell growth,mitochondrial oxidative phosphorylation,lipid metabolism,and cardiac disease,and its mutation can lead to alterations in protein interaction networks.
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