As part of my PhD project, I have spent my Secondment period in Sweden in the laboratory of Maria Falkenberg at the University of Gothenburg. I worked in Falkenberg’s lab from 5th to 23th November 2018 and from 1^st to 12^th April 2019 aiming to characterize the in vitro function of POLγA variants.

POLγA is the catalytic subunit of POLγ, responsible for replicating mitochondrial DNA (mtDNA). DNA mutations affecting POLγA are a major cause of human mitochondrial disease and are associated with a continuum spectrum of disease manifestations. My PhD project is focused in the characterization of one of the most common mutations which determines a p.A467T substitution in POLγA. To gain insights into complex clinical and molecular pathophysiology of POLG-related diseases, my current lab generated a POLγA^A449T knock-in mouse model, corresponding to the p.A467T substitution in humans. While the characterization of this mouse model is undergoing, I decided to further characterize POLγA^A449T protein in vitro and also compare it with the corresponding p.A467T substitution in humans (in Falkenberg’s lab).

Falkenberg’s lab is recognized as one of the leading experts in mitochondrial DNA replication and have developed over the years several in vitro assays to characterize several proteins involved in this process, including POLγA. During my Secondment in Falkenberg’s lab I was able to learn several techniques and access in vitro the effects of pathogenic variants in the diverse functions of POLγA. These include: DNA polymerase and exonuclease activities; binding affinity to PolγB subunits; binding affinity to a DNA template; and protein thermostability.Furthermore, I was able to compare the diverse functions between mouse POLγA and human POLγA. This data was of great value to further support the transgenic mouse as a relevant model for POLγA-related diseases.

An additional visit to Falkenberg’s lab is expected towards the end of my PhD project, to complete some experiments and to continue our collaboration.