Metabolome and Proteome Profiling of Complex I Deficiency Induced by Rotenone

Complex I (CI; NADH dehydrogenase) deficiency causes mitochondrial diseases, including Leigh syndrome. A variety of clinical symptoms of CI deficiency are known, including neurodegeneration. Here, we report an integrative study combining liquid chromatography–mass spectrometry (LC–MS)-based metabolome and proteome profiling in CI deficient HeLa cells. We report a rapid LC–MS-based method for the relative quantification of targeted metabolome profiling with an additional layer of confidence by applying multiple reaction monitoring (MRM) ion ratios for further identity confirmation and robustness. The proteome was analyzed by label-free quantification (LFQ). More than 6000 protein groups were identified. Pathway and network analyses revealed that the respiratory chain was highly deregulated, with metabolites such as FMN, FAD, NAD⁺, and ADP, direct players of the OXPHOS system, and metabolites of the TCA cycle decreased up to 100-fold. Synthesis of functional iron–sulfur clusters, which are of central importance for the electron transfer chain, and degradation products like bilirubin were also significantly reduced. Glutathione metabolism on the pathway level, as well as individual metabolite components such as NADPH, glutathione (GSH), and oxidized glutathione (GSSG), was downregulated. Overall, metabolome and proteome profiles in CI deficient cells correlated well, supporting our integrated approach

Integrative Analysis of Transcriptomics, Proteomics, and Metabolomics Data of White Adipose and Liver Tissue of High-Fat Diet and Rosiglitazone-Treated Insulin-Resistant Mice Identified Pathway Alterations and Molecular Hubs

The incidences of obesity and type 2 diabetes are rapidly increasing and have evolved into a global epidemic. In this study, we analyzed the molecular effects of high-fat diet (HFD)-induced insulin-resistance on mice in two metabolic target tissues, the white adipose tissue (WAT) and the liver. Additionally, we analyzed the effects of drug treatment using the specific PPARγ ligand rosiglitazone. We integrated transcriptome, proteome, and metabolome data sets for a combined holistic view of molecular mechanisms in type 2 diabetes. Using network and pathway analyses, we identified hub proteins such as SDHB and SUCLG1 in WAT and deregulation of major metabolic pathways in the insulin-resistant state, including the TCA cycle, oxidative phosphorylation, and branched chain amino acid metabolism. Rosiglitazone treatment resulted mainly in modulation via PPAR signaling and oxidative phosphorylation in WAT only. Interestingly, in HFD liver, we could observe a decrease of proteins involved in vitamin B metabolism such as PDXDC1 and DHFR and the according metabolites. Furthermore, we could identify sphingosine (Sph) and sphingosine 1-phosphate (SP1) as a drug-specific marker pair in the liver. In summary, our data indicate physiological plasticity gained by interconnected molecular pathways to counteract metabolic dysregulation due to high calorie intake and drug treatment

Quantitative Global Proteomics of Yeast PBP1 Deletion Mutants and Their Stress Responses Identifies Glucose Metabolism, Mitochondrial, and Stress Granule Changes

The yeast protein PBP1 is implicated in very diverse pathways. Intriguingly, its deletion mitigates the toxicity of human neurodegeneration factors. Here, we performed label-free quantitative global proteomics to identify crucial downstream factors, either without stress or under cell stress conditions (heat and NaN₃). Compared to the wildtype BY4741 strain, PBP1 deletion always triggered downregulation of the key bioenergetics enzyme KGD2 and the prion protein RNQ1 as well as upregulation of the leucine biosynthesis enzyme LEU1. Without stress, enrichment of stress response factors was consistently detected for both deletion mutants; upon stress, these factors were more pronounced. The selective analysis of components of stress granules and P-bodies revealed a prominent downregulation of GIS2. Our yeast data are in good agreement with a global proteomics and metabolomics publication that the PBP1 ortholog ATAXIN-2 (ATXN2) knockout (KO) in mouse results in mitochondrial deficits in leucine/fatty acid catabolism and bioenergetics, with an obesity phenotype. Furthermore, our data provide the completely novel insight that PBP1 mutations in stress periods involve GIS2, a plausible scenario in view of previous data that both PBP1 and GIS2 relocalize from ribosomes to stress granules, interact with poly­(A)-binding protein in translation regulation and prevent mitochondrial precursor overaccumulation stress (mPOS). This may be relevant for human diseases like spinocerebellar ataxias, amyotrophic lateral sclerosis, and the metabolic syndrome

Additional file 11: Fig. S4. of Progression of pathology in PINK1-deficient mouse brain from splicing via ubiquitination, ER stress, and mitophagy changes to neuroinflammation

Transcriptional response of innate immunity factors to 24 h treatment with uncoupling drug FCCP and subsequent mitophagy, in dependence on PINK1. Three independent experiments in SH-SY5Y human neuroblastoma (above) and murine embryonal fibroblast cells (below) documented the expression of key inflammatory factors in untreated versus drug-treated cells, comparing control with PINK1-deficiency. The bar graphs show mean and standard error of the mean, illustrating the significance with asterisks (* p < 0.05, ** p < 0.01, *** p < 0.001). (TIFF 538 kb