title: Zebrafish lacking functional DNA polymerase gamma survive to juvenile stage, despite rapid and sustained mitochondrial DNA depletion, altered energetics and growth.
abstract: DNA polymerase gamma (POLG) is essential for replication and repair of mitochondrial DNA (mtDNA). Mutations in POLG cause mtDNA instability and a diverse range of poorly understood human diseases. Here, we created a unique Polg animal model, by modifying polg within the critical and highly conserved polymerase domain in zebrafish. polg(+/-) offspring were indistinguishable from WT siblings in multiple phenotypic and biochemical measures. However, polg(-/-) mutants developed severe mtDNA depletion by one week post-fertilization (wpf), developed slowly and had regenerative defects, yet surprisingly survived up to 4 wpf. An in vivo mtDNA polymerase activity assay utilizing ethidium bromide (EtBr) to deplete mtDNA, showed that polg(+/-) and WT zebrafish fully recover mtDNA content two weeks post-EtBr removal. EtBr further reduced already low levels of mtDNA in polg(-/-) animals, but mtDNA content did not recover following release from EtBr. Despite significantly decreased respiration that corresponded with tissue-specific levels of mtDNA, polg(-/-) animals had WT levels of ATP and no increase in lactate. This zebrafish model of mitochondrial disease now provides unique opportunities for studying mtDNA instability from multiple angles, as polg(-/-) mutants can survive to juvenile stage, rather than lose viability in embryogenesis as seen in Polg mutant mice.
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA.
authors.forename: Jennifer J
authors.initials: JJ
authors.lastname: Rahn
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA.
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Bestman
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA.
authors.forename: Krista D
authors.initials: KD
authors.lastname: Stackley
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, SC 29425, USA chans@musc.edu.
authors.forename: Sherine S L
authors.initials: SS
authors.lastname: Chan
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D058500:Animal Fins / Q000502:physiology; D000818:Animals; D000074002:DNA Polymerase gamma; D004272:DNA, Mitochondrial / Q000032:analysis*; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005818:Genetic Engineering; D006019:Glycolysis; D023421:Models, Animal; D009154:Mutation; D010101:Oxygen Consumption; D012038:Regeneration; D016019:Survival Analysis; D015027:Zebrafish / Q000235:genetics* / Q000254:growth & development; D029961:Zebrafish Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
chemical_list: D004272:DNA, Mitochondrial; D029961:Zebrafish Proteins; D000255:Adenosine Triphosphate; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase