title: CoQ10 Deficiency: The Lysosomal Paradigm
Creators: name: Heaton, R
Publisher: name: Liverpool John Moores University
Year: 2022

description: Coenzyme Q10 (CoQ10) deficiency currently represents the only treatable disorder of the mitochondrial electron transport chain (METC). Generally, in patients with CoQ10 deficiency there is some form of neurological impairment typically presenting as either ataxia and/or seizures. Currently the preponderance of research relating to a CoQ10 deficiency has focused on its effect on mitochondrial function, and there is a paucity of information on how this may affect other organelles. Interestingly, the lysosome has been found to have a large concentration of CoQ10 localised in its membrane which is thought to play a role in the normal acidification of the lysosomal lumen. In order to investigate the effect of a CoQ10 deficiency on lysosomal acidification, we established a neuronal cell model of CoQ10 deficiency via the treatment of the cell line SH-SY5Y with Para-AminoBenzoic Acid (PABA) a competitive inhibitor of the CoQ10 biosynthetic enzyme, para-hydroxybenzoate— polyprenyltransferase (COQ2). A single 1 mM (5 days) treatment with PABA resulted in a significant decrease of up to 58% in cellular CoQ10 status (p &lt; 0.05). This decrease in cellular CoQ10 status was then found to be associated with a significant decrease in both the LysoSensor (23%) and LysoTracker 167 (35%) fluorescence; probes used to measure lysosomal lumen pH (p &lt; 0.05). This suggests that the lysosomal pH has been significantly increased as the result of a deficit in CoQ10 status. Additionally, a third pH probe was used, LysoSensor 160, which was able to accurately determine the pH of the lysosomal lumen. It was found that a decrease in neuronal CoQ10 status of (58%) resulted in a significant increase in lysosomal pH from 4.1 in control cells, to 6.3 in the CoQ10 deficient state. This finding provides evidence of the vulnerability of the lysosomal pH to changes in CoQ10 status and is the first study of its kind to investigate this phenomenon. Subsequently, the neuronal cell model was utilised to evaluate whether the observed increase in lysosomal pH associated with a CoQ10 deficiency and could it be reversed 3 via supplementation with CoQ10 at levels reported to improve mitochondrial function in patient fibroblast studies. Treatment with CoQ10 (5 μM, 3 days) was found to restore cellular CoQ10 concentration above controls (p &lt; 0.005) and this was associated with an increase in both LysoTracker and LysoSensor 167 fluorescence to around 90% of control levels (p &lt; 0.05), suggesting that lysosomal pH had been restored to 90% of its original level. Analysis of the LysoSensor 160 probe showed CoQ10 supplementation was associated with a reduction in lysosomal pH from 6.3 to around 4.7. In addition to the increase in lysosomal pH, a number of other cellular aspects were also investigated in an attempt to determine some possible mechanisms causing the increase in lysosomal pH in CoQ10 deficient neuronal cells. General METC function was measured using the fluorescent probes JC-1 and MTG together with measurements of the mitochondrial marker enzyme, citrate synthase (CS) and assessments of total cellular ATP concentration. JC-1 is a specific probe which measures mitochondrial membrane potential (ΔΨm) and using this probe it was found that there was no significant difference in the fluorescence between the control and CoQ10 deficient cells. Additionally, MTG, a probe that assesses mitochondrial mass also found no significant difference between control cells and CoQ10 deficient cells. In contrast, CS activity and cellular ATP concentration were found to be significantly different between control and CoQ10 deficient neurones. CS activity was significantly increased (p&lt; 0.005) in the CoQ10 deficient neurones contradicting the MTG result. In addition, cellular ATP concentration was found to be significantly decreased (15% lower than controls) in the CoQ10 deficient neurones. The deficit in cellular ATP status may impact upon the lysosomal pH given the requirement of the lysosomal V-ATPase for cytosolic ATP. In this study we also measured total cellular reactive oxygen species (ROS) levels due to because of CoQ10’s role as a potent antioxidant and the lysosomes sensitivity to oxidative stress induced impairment. Commensurate with the deficit in neuronal cellular CoQ10 status a significant increase in cellular ROS levels (29% p&lt; 0.005) was determined which could also contribute to lysosomal alkalisation.