Models of mitochondrial dysfunction with inducible expression of Polg pathogenic mutant variant

Kubekina MV1, Kalinina AA2, Korshunova DS1, Bruter AV1, Silaeva YY1
About authors

1 Institute of Gene Biology, Moscow, Russia

2 Blokhin Russian Cancer Research Center, Moscow, Russia

Correspondence should be addressed: Marina V. Kubekina
Beskudnikovsky bulvar, 32, korpus 1, Moscow, 127474, Russia; moc.liamg@ymukyram

About paper

Funding: the study was supported by Russian Foundation for Basic Research, RFBR Project № 19-34-90073.

Author contribution: Kubekina MV — literature analysis, experimental research, data analysis and interpretation, oligo design, manuscript writing; Kalinina AA, Korshunova DS — experimental research; Bruter AV — literature analysis, research planning, data analysis and interpretation; Silaeva YY — literature analysis, research planning, data analysis and interpretation, scientific editing of the manuscript.

Compliance with ethical standards: the study was approved by Ethical Review board at the Institute of Gene Biology (Protocol of 05 December 2021) and carried out in strict compliance with the Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes.

Received: 2022-04-14 Accepted: 2022-04-28 Published online: 2022-04-27

Mitochondrial dysfunctions, which underlie many systemic diseases in animals and humans, may arise from accumulation of mutations in the mitochondrial genome. PolG-alpha enzyme encoded by Polg gene is crucial for replication and repair of the mitochondrial genome. The aim of this study was to assess the possible role of Polg mutations in mitochondrial dysfunctions using in vitro and in vivo animal models. The experiments involved transgenic mice with inducible expression of Polg mutant variant; the methods included cell culture, real time PCR assay, fluorescence flow cytometry, and skeletal muscle functional tests. The results indicate that mouse embryonic fibroblasts (MEFs) expressing Polg pathogenic mutant variant have decreased mitochondrial membrane potential and increased expression of mitophagy markers compared with control cultures. Transgenic animals with systemic expression of the pathogenic variant develop mitochondrial dysfunction which significantly affects muscular performance. In addition, systemic expression of mutated Polg in transgenic animals significantly inhibits expression of TCR subunit α and CD3 coreceptor complex subunits δ and ε in total splenocyte populations and significantly affects cellularity of the thymus without altering its CD4/CD8 subpopulation ratio. Thus, inducible expression of mutated Polg in transgenic animals provides a relevant model for studying mitochondrial dysfunction and its treatment in vitro and in vivo.

Keywords: genome editing, mitochondrial dysfunction, PolG-alpha enzyme, transgenic animals, mitochondrial membrane potential, mitophagy