Dysregulated Provision of Oxidisable Substrates to the Mitochondria in ME/CFS Lymphoblasts

Abstract

Although understanding of the biomedical basis of Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is growing, the underlying pathological mechanisms remain uncertain. We recently reported a reduction in the proportion of basal oxygen consumption due to ATP synthesis by Complex V in ME/CFS patient-derived lymphoblast cell lines, suggesting mitochondrial respiratory inefficiency. This was accompanied by elevated respiratory capacity, elevated mammalian target of rapamycin complex 1 (mTORC1) signaling activity and elevated expression of enzymes involved in the TCA cycle, fatty acid β-oxidation and mitochondrial transport. These and other observations led us to hypothesise the dysregulation of pathways providing the mitochondria with oxidisable substrates. In our current study, we aimed to revisit this hypothesis by applying a combination of whole-cell transcriptomics, proteomics and energy stress signaling activity measures using subsets of up to 34 ME/CFS and 31 healthy control lymphoblast cell lines from our growing library. While levels of glycolytic enzymes were unchanged in accordance with our previous observations of unaltered glycolytic rates, the whole-cell proteomes of ME/CFS lymphoblasts contained elevated levels of enzymes involved in the TCA cycle (p = 1.03 × 10−4), the pentose phosphate pathway (p = 0.034, G6PD p = 5.5 × 10−4), mitochondrial fatty acid β-oxidation (p = 9.2 × 10−3), and degradation of amino acids including glutamine/glutamate (GLS p = 0.034, GLUD1 p = 0.048, GOT2 p = 0.026), branched-chain amino acids (BCKDHA p = 0.028, BCKDHB p = 0.031) and essential amino acids (FAH p = 0.036, GCDH p = 0.006). The activity of the major cellular energy stress sensor, AMPK, was elevated but the increase did not reach statistical significance. The results suggest that ME/CFS metabolism is dysregulated such that alternatives to glycolysis are more heavily utilised than in controls to provide the mitochondria with oxidisable substrates.

Bioenergetic and Proteomic Profiling of Immune Cells in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome Patients: An Exploratory Study

“…We observed, however, lower levels of ATP5F1E, a complex V subunit. Previous studies in immortalised lymphocytes from ME/CFS patients have shown complex V deficiency and hyperactivation of mTORC1 [31,70]. In addition, downregulation of SLC25A24 can also indicate disturbed ATP synthesis [71].…”

Lymphoblastoid Cell Lines as Models to Study Mitochondrial Function in Neurological Disorders

“…The examples provided here suggest that these cells will be useful for the development of diagnostic tests (which in most cases do not exist), identification of drug targets and testing of pharmacological agents, and are a worthwhile model for studying mitochondrial function in neurological disorders. [44,59]…”

“…Despite these obvious differences, the methylation pattern and gene expression profiles of lymphocytes are largely maintained in LCLs and display many of the specific characteristics of primary B cells [ 58 ]. This is also true for many diseases, and clear differences in the transcriptomes and the epigenomes of patients and controls have been identified in LCLs for numerous diseases [ 59 , 60 , 61 , 62 , 63 ]. LCLs are increasingly being used as models of disease and given their high oxidative nature, are ideal systems for studying mitochondrial function.…”

“…TORC1 is able to activate transcription factors and separately upregulate the translation of many proteins, including mitochondrial proteins. All five mitochondrial respiratory complexes were indeed upregulated, as were enzymes involved in the TCA cycle, fatty acid uptake, β-oxidation and other pathways that serve as alternatives to glycolysis involved in provisioning the mitochondria with oxidizable substrates [ 59 ]. This elevation of expression may represent a cellular “attempt” to compensate for the inefficiency of Complex V. The upregulation of mitochondrial proteins was also reported in non-immortalised lymphocytes and saliva, supporting the utility of LCLs as a model system for this disease [ 60 , 118 , 119 ].…”

Chronic Activation of AMPK Induces Mitochondrial Biogenesis through Differential Phosphorylation and Abundance of Mitochondrial Proteins in Dictyostelium discoideum

“…This could combine with AMPK’s direct inhibitory phosphorylation of the RAPTOR subunit of mTORC1 to inhibit its activity and produce a decrease in cell growth and division [ 97 ]. mTORC1 is known for stimulation of expression of OXPHOS proteins [ 98 , 99 ]; hence, it is likely that the downregulation of OXPHOS components in AMPKα-overexpressing Dictyostelium cells results from the inhibition of mTORC1 activity. Enhanced mitochondrial biogenesis is induced by as yet unidentified mitochondrial protein targets, which, upon direct or indirect phosphorylation by AMPK, leads to dramatic changes in the abundance and/or phosphorylation-dependent activity of mitochondrial proteins involved in energy metabolism, protein synthesis, and membrane biogenesis.…”