TIMMDC1/C3orf1 Functions as a Membrane-Embedded Mitochondrial Complex I Assembly Factor through Association with the MCIA Complex

Abstract

Complex I (CI) of the electron transport chain, a large membrane-embedded NADH dehydrogenase, couples electron transfer to the release of protons into the mitochondrial inner membrane space to promote ATP production through ATP synthase. In addition to being a central conduit for ATP production, CI activity has been linked to neurodegenerative disorders, including Parkinson's disease. CI is built in a stepwise fashion through the actions of several assembly factors. We employed interaction proteomics to interrogate the molecular associations of 15 core subunits and assembly factors previously linked to human CI deficiency, resulting in a network of 101 proteins and 335 interactions (edges). TIMMDC1, a predicted 4-pass membrane protein, reciprocally associated with multiple members of the MCIA CI assembly factor complex and core CI subunits and was localized in the mitochondrial inner membrane, and its depletion resulted in reduced CI activity and cellular respiration. Quantitative proteomics demonstrated a role for TIMMDC1 in assembly of membrane-embedded and soluble arms of the complex. This study defines a new membrane-embedded CI assembly factor and provides a resource for further analysis of CI biology.

Regulation of Mitochondrial Complex I Biogenesis in Drosophila Flight Muscles

“…However, in stark contrast to the other CI subunits, we consistently observed (in 6 biological replicates taken at different time points of the day to control for circadian regulation) that dNDUFA10 was downregulated in the mhc>dNDUFS5 RNAi sample; indicating that incorporation of dNDUFS5 into CI is necessary to stabilize or promote incorporation of dNDUFA10 into the complex (Figure 5D). In mammalian systems, at least five CI assembly factors – ECSIT, TMEM126B, NDUFAF1, ACAD9 and TIMMDC1 – are typically found associated with CI assembly intermediates, and have been dubbed the M itochondrial C omplex I A ssembly (MCIA) complex (Guarani et al, 2014; Heide et al, 2012; Nouws et al, 2010; Vogel et al, 2007). We found four of these assembly factors (dECSIT, dNDUFAF1, dACAD9 and dTIMMDC1), associated with the 700 kDa assembly intermediate that were upregulated in the mhc>dNDUFS5 RNAi samples, further confirming that it is a true assembly intermediate in CI biogenesis (Figure 5D and Table S4).…”

“… (A) Schematic representation of how the 44 different subunits of bovine CI are arranged to produce the L-shaped topology; adapted from (Guarani et al, 2014). The asterisk denotes subunits for which an ortholog was not identified in Drosophila by DIOPT.…”

Biallelic Mutations in TMEM126B Cause Severe Complex I Deficiency with a Variable Clinical Phenotype

“…10 Complexome profiling revealed TMEM126B to be a component of the mitochondrial complex I assembly (MCIA) complex alongside proteins ACAD9, ECSIT, NDUFAF1, and TIMMDC1, thus establishing TMEM126B (MIM: 615533) as a candidate gene for complex I deficiency. 10,11 With access to subjects harboring putative TMEM126B defects, we provide functional evidence to support the pathogenicity of these TMEM126B variants, unequivocally establishing this gene as a cause of complex I deficiency in association with either a severe multisystem presentation in infancy or pure myopathy in later child-or adulthood. This report describes the clinical, biochemical, and molecular findings in six cases of TMEM126B-related mitochondrial disease and validates the importance of proteomic approaches in identifying disease-associated genes whose physiological roles have been previously undetermined.…”

Mutations in Complex I Assembly Factor TMEM126B Result in Muscle Weakness and Isolated Complex I Deficiency

“…Interestingly, NDUFAF1 and ACAD9, assembly factors acting at the level of the CI membrane arm where TMEM126B is expected to interact, 25,29,32 showed increased steady-state levels in protein extracts from subject 1 fibroblasts. In fibroblasts of subjects 2 and 3, CI subunits NDUFS3, NDUFB8, and NDUFA9 were decreased, which is in line with the decreased assembly of complex I (Figures 3D and 3E).…”

“…His height and weight were 1.49 m and 37.7 kg, respectively. Metabolic investigations showed permanent hyperlactacidemia (2.8-7.4 mmol/L), elevated lactate/pyruvate ratios (21)(22)(23)(24)(25)(26)(27)(28)(29)(30)(31)(32), hyperalaninemia (702 mmol/L; normal range ¼ 274 5 50 mmol/L), and elevated lactate levels in urine (2,775 mmol/mmol creatinine; control values < 84 mmol/mmol creatinine). Ophthalmological and cardiac examination was normal.…”