Investigating the adaptive mitochondrial shuttles and metabolic reprogramming of transporters in complex I (Ndufs4) knockout mice

Abstract

Mitochondrial disease is one of the most prevalent inherited paediatric disorders among children, with a prevalence of 1 in 5000 children. Leigh syndrome is the result of a Complex I (CI) deficiency and disrupts the redox balance needed for the function of various dehydrogenase enzymes. Due to the extensive heterogeneity of the disease, it remains a challenge to diagnose and treat mitochondrial diseases. One of the most overlooked areas for possible treatment is the solute carriers of the inner mitochondrial membrane (IMM). These solute carriers function together to form shuttle systems that transport electrons over the impermeable IMM, which can possibly aid in recovery of the disrupted redox balance. Targeted transcriptomic analysis was done on liver, heart and brain tissue collected from a CI deficient (Ndufs4 knockout) mice model to analyse the gene expression of the selected solute carriers part of the malate-aspartate shuttle and the citrate-pyruvate shuttle. The proteins of the glycerol-3-phosphate shuttle was also included due to its ability to also carry electrons over the IMM. This was done by making use of the Ion GeneStudio™ S5 Semiconductor Sequencer accompanied by the Ion Chef™ Instrument. Targeted metabolomics was also done on the selected tissue on the metabolites transported by the solute carriers, also to deduce if there are any changes in their abundance with diseases like Leigh syndrome. This was done via GC-TOF-MS and LC-MS/MS analysis. Transcriptomic analysis revealed for the liver tissue, Gpd1 (glycerol-3-phosphate dehydrogenase 1), Mdh1 (malate dehydrogenase 1), Me1 (malic enzyme 1) and Slc25a1 (solute carrier family 25 member 1, citrate carrier) were all down regulated in the knockout group. For the heart tissue, Slc25a22 (solute carrier family 25 member 22, glutamate carrier) and Me1 (malic enzyme 1) were also down regulated in the knockout group and for the brain tissue, Me3 (malic enzyme 3), Slc25a3 (solute carrier 25 member 3, phosphate carrier) and Slc25a22 (solute carrier family 25 member 22, glutamate carrier) were all up regulated in the knockout group. Changes in the expression of these genes all seem to be an effect of the redox imbalance due to complex I deficiency or revolve around an attempt to maintain energy production. Metabolomic analysis indicated an increase in abundance in metabolites associated with the down-regulated enzymes and shuttles. These changes all seem to be an effect of the complex I knockout and not an attempt to compensate for the disruption of the cells function. Considering this study involved an unconventional method for gene expression analysis, the overall

success in the outcome of this study can be attributed to the combination of transcriptomics and metabolomics data to give a better understanding of the events at molecular level with abnormalities such as complex I deficiency.