Z HOT TOPIC Z 

Author Block: Heidi Britton, Bryce Pasqualotto, Leonard Foster, Gordon Rintoul, Claire Sheldon
Author Disclosure Block: H. Britton: None. B. Pasqualotto: None. L. Foster: None. G. Rintoul: None. C. Sheldon: None.

Abstract Body:

Purpose: LHON is primarily associated with one of three point mutations in the mitochondrial DNA encoding NADH-ubiquinone oxidoreductase subunits 1, 4, or 6. However, these mutations are necessary but not sufficient for LHON to occur and both genetic and physiological conditions affect the onset and severity of LHON. As a result, analysis of the proteome is of significant interest to the field, however very limited data is available to the field (e.g. no entries in the ProteomeXchange or PRIDE databases). 

Methods: In this pilot study, skin punch biopsies were obtained from a subject with vision loss, expressing the 11778 mutation of LHON, in addition to an age- and sex-matched control. Cultures were maintained in either glucose- or galactose-containing media, to assess the influence of metabolic conditions requiring exclusive use of oxidative phosphorylation. Liquid chromatography-tandem mass spectrometry, incorporating stable isotope labeling for quantitation, was used to compare the proteomic profiles of cells under these conditions. Data was analyzed with MaxQuant, a quantitative proteomics data package designed for large-scale MS data sets. All proteins with a ratio of LHON to control of ≥2-fold over- or under-expressed, or with a p-value < 0.05 and a minimum of 3 replicates, were further analyzed against the MitoMiner 4.0, neXtProt, and Open Targets Target Validation databases to assess mitochondrial association and function. 

Results: 102 under-expressed and 115 over-expressed proteins were identified, of which 20 and 23 were found to be associated with the mitochondria, respectively. Under-expressed mitochondrial proteins included those associated with the electron transport chain, protein trafficking, oxidative stress, and the inner mitochondrial membrane. Over-expressed mitochondrial proteins were associated with mitochondrial protein synthesis and oxidative stress. 

Conclusions: Understanding the proteome is important to identifying factors that result in symptomatic LHON, and thereby important to finding treatments to delay or prevent symptom onset. Our findings support the conventional models of pathogenesis in LHON involving impaired energy metabolism and ROS imbalance and identify several other potential targets for future study.