Year: 2018
Type: article
Abstract: Polynucleotide phosphorylase (PNPase) is an essential mitochondria-localized exoribonuclease implicated in multiple biological processes and human disorders. To reveal role(s) for PNPase in mitochondria, we established PNPase knockout (PKO) systems by first shifting culture conditions to enable cell growth with defective respiration. Interestingly, PKO established in mouse embryonic fibroblasts (MEFs) resulted in the loss of mitochondrial DNA (mtDNA). The transcriptional profile of PKO cells was similar to rho0 mtDNA deleted cells, with perturbations in cholesterol (FDR = 6.35 x 10-13), lipid (FDR = 3.21 x 10-11), and secondary alcohol (FDR = 1.04x10-12) metabolic pathway gene expression compared to wild type parental (TM6) MEFs. Transcriptome analysis indicates processes related to axonogenesis (FDR = 4.49 x 10-3), axon development (FDR = 4.74 x 10-3), and axonal guidance (FDR = 4.74 x 10-3) were overrepresented in PKO cells, consistent with previous studies detailing causative PNPase mutations in delayed myelination, hearing loss, encephalomyopathy, and chorioretinal defects in humans. Overrepresentation analysis revealed alterations in metabolic pathways in both PKO and rho0 cells. Therefore, we assessed the correlation of genes implicated in cell cycle progression and total metabolism and observed a strong positive correlation between PKO cells and rho0 MEFs compared to TM6 MEFs. We quantified the normalized biomass accumulation rate of PKO clones at 1.7% (SD ± 2.0%) and 2.4% (SD ± 1.6%) per hour, which was lower than TM6 cells at 3.3% (SD ± 3.5%) per hour. Furthermore, PKO in mouse inner ear hair cells caused progressive hearing loss that parallels human familial hearing loss previously linked to mutations in PNPase. Combined, our study reports that knockout of a mitochondrial nuclease results in mtDNA loss and suggests that mtDNA maintenance could provide a unifying connection for the large number of biological activities reported for PNPase.
Source: PLoS ONE
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,382
Funders: California Institute for Regenerative Medicine, California Institute of Regenerative Medicine, National Institutes of Health, National Cancer Institute, National Institute on Deafness and Other Communication Disorders, National Institute of General Medical Sciences, National Institute of Neurological Disorders and Stroke, Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, University of California Los Angeles, Air Force Office of Scientific Research
Awards: (PQD5) Mass Profiling Melanoma Responses to Improve Therapy Choices and Prognosis, G1820511087, RNA trafficking in mitochondria, Reverse Mitochondrial Genetics Enabled by Blast, G4754645965, RNA Import into Mitochondria and Neurodegenerative Disorders, G6318647045, CRTC2 in Cellular Development, Function, and Neoplasia, Research Tranining in Cellular and Molecular Biology, A Fourth Outcome: DNA Damage and the Differentiation of B Cells, A small molecule tool for reducing the malignant potential in reprogramming human iPSCs and ESCs, G9000269057, Cytoskeletal stability in stereocilia maintenance, BIOGENESIS OF THE MITOCHONDRIAL INNER MEMBRANE, UCLA Tumor Immunology Institutional Training Grant
OpenAlex ID: W2883517429