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PNPase knockout results in mtDNA loss and an altered metabolic gene expression program.
- Source :
-
PloS one [PLoS One] 2018 Jul 19; Vol. 13 (7), pp. e0200925. Date of Electronic Publication: 2018 Jul 19 (Print Publication: 2018). - Publication Year :
- 2018
-
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.<br />Competing Interests: The authors have declared that no competing interests exist.
- Subjects :
- Animals
Cell Cycle
DNA, Mitochondrial genetics
Female
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
Mitochondria genetics
Mutation
Polyribonucleotide Nucleotidyltransferase genetics
Basic Helix-Loop-Helix Transcription Factors physiology
DNA, Mitochondrial metabolism
Gene Expression Regulation
Hearing Loss physiopathology
Mitochondria metabolism
Polyribonucleotide Nucleotidyltransferase metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1932-6203
- Volume :
- 13
- Issue :
- 7
- Database :
- MEDLINE
- Journal :
- PloS one
- Publication Type :
- Academic Journal
- Accession number :
- 30024931
- Full Text :
- https://doi.org/10.1371/journal.pone.0200925