TAT-Conjugated NDUFS8 Can Be Transduced into Mitochondria in a Membrane-Potential-Independent Manner and Rescue Complex I Deficiency

NADH dehydrogenase (ubiquinone) Fe-S protein 8 (NDUFS8) is a nuclear-encoded core subunit of human mitochondrial complex I. Defects in NDUFS8 are associated with Leigh syndrome and encephalomyopathy. Cell-penetrating peptide derived from the HIV-1 transactivator of transcription protein (TAT) has been successfully applied as a carrier to bring fusion proteins into cells without compromising the biological function of the cargoes. In this study, we developed a TAT-mediated protein transduction system to rescue complex I deficiency caused by NDUFS8 defects. Two fusion proteins (TAT-NDUFS8 and NDUFS8-TAT) were exogenously expressed and purified from Escherichia coli for transduction of human cells. In addition, similar constructs were generated and used in transfection studies for comparison. The results showed that both exogenous TAT-NDUFS8 and NDUFS8-TAT were delivered into mitochondria and correctly processed. Interestingly, the mitochondrial import of TAT-containing NDUFS8 was independent of mitochondrial membrane potential. Treatment with TAT-NDUFS8 not only significantly improved the assembly of complex I in an NDUFS8-deficient cell line, but also partially rescued complex I functions both in the in-gel activity assay and the oxygen consumption assay. Our current findings suggest the considerable potential of applying the TAT-mediated protein transduction system for treatment of complex I deficiency.

Mitochondrial Complex I Deficiency among Egyptian Pediatric Patients with Steroid-Resistant Nephrotic Syndrome

Background and objectives. Nephrotic Syndrome (NS) is one of the most common glomerular diseases among children. Up to 20% of patients are steroid resistant (SRNS) representing a challenging subset at high risk of developing end-stage renal disease. Renal manifestations of mitochondrial diseases (MIDs) include nephrotic syndrome, renal insufficiency, nephrolithiasis, Bartter-like syndrome, focal segmental glomerulosclerosis, and nephrocalcinosis. The objective of the current study is to measure the activity of mitochondrial complex I in renal biopsies obtained from pediatric patients diagnosed with SRNS compared to steroid-sensitive nephrotic syndrome (SSNS) patients in order to elucidate its role in pathogenesis and the prognosis for further genetic work. Subjects and Method. Renal biopsies of 120 patients diagnosed with nephrotic syndrome based on clinical and laboratory findings, divided into two groups, SRNS (60 patients) and SSNS (60 patients). Pathological study and spectrophotometric measurement of mitochondrial complex I in renal biopsy and muscle homogenates were performed for both groups. Results. Positive consanguinity was a remarkable finding in 44 patients among the SRNS group (73%), compared with 33 patients among the SSNS group (55%). Complex I activity was significantly lower in the SRNS group (0.2657 ± 0.1831 nmol/ml/min), than in the SSNS group (0.4773 ± 0.1290 nmol/ml/min) ( p < 0.001 ). There was a significant positive correlation between complex I activity and the heaviness of proteinuria among the SRNS group (r 0.344, p < 0.001 ). There were statistically significant differences in serum C3 and C4 levels between both groups ( p < 0.001 , 0.053, respectively). Conclusion. Mitochondrial complex I deficiency in patients who have a nephrotic syndrome complaint may play a role in their responsiveness to steroid therapy and the development of SRNS and even the prognosis of their illness.

The mitochondrial calcium uniporter compensates for Complex I dysfunction

Calcium (Ca2+) entering mitochondria potently stimulates ATP synthesis. Increases in Ca2+ preserve energy synthesis in cardiomyopathies caused by mitochondrial dysfunction, and occur due to enhanced activity of the mitochondrial Ca2+ uniporter channel. The signaling mechanism that mediates this compensatory increase remains unknown. Here, we find that increases in the uniporter are due to impairment in Complex I of the electron transport chain (ETC). In normal physiology, Complex I promotes uniporter degradation via an interaction with the uniporter pore-forming subunit, a process we term Complex I-induced protein turnover (CLIPT). When Complex I dysfunction ensues, contact with the uniporter is inhibited, preventing degradation, and leading to a build-up in functional channels. Preventing uniporter activity leads to early demise in Complex I-deficient animals. Conversely, enhancing uniporter stability rescues survival and function in Complex I deficiency. Taken together, our data identify a fundamental pathway producing compensatory increases in Ca2+ influx during Complex I impairment.

Optic atrophy–associated TMEM126A is an assembly factor for the ND4-module of mitochondrial complex I

Mitochondrial disease is a debilitating condition with a diverse genetic etiology. Here, we report that TMEM126A, a protein that is mutated in patients with autosomal-recessive optic atrophy, participates directly in the assembly of mitochondrial complex I. Using a combination of genome editing, interaction studies, and quantitative proteomics, we find that loss of TMEM126A results in an isolated complex I deficiency and that TMEM126A interacts with a number of complex I subunits and assembly factors. Pulse-labeling interaction studies reveal that TMEM126A associates with the newly synthesized mitochondrial DNA (mtDNA)-encoded ND4 subunit of complex I. Our findings indicate that TMEM126A is involved in the assembly of the ND4 distal membrane module of complex I. In addition, we find that the function of TMEM126A is distinct from its paralogue TMEM126B, which acts in assembly of the ND2-module of complex I.

Degradation of mitochondrial structure and deficiency of complex I were associated with the transgenic CMS of rice

Background Mitochondria play a significant role in plant cytoplasmic male sterility (CMS). In our previous study, mitochondrial complex I genes, nad4, nad5, and nad7 showed polymorphisms between the transgenic CMS line M2BS and its wild type M2B. The sterility mechanism of the M2BS at cytological, physiological, biochemical, and molecular level is not clear. Results Cytological observation showed that the anthers were light yellow, fissured, invalid in KI-I2, and full of irregularly typical abortion pollen grains in M2BS. Transmission electron microscopic (TEM) observation revealed no nucleus and degraded mitochondria with obscure cristae in anther cells of M2BS. The results of staining for H2O2 presented a large number of electron dense precipitates (edp) in intercellular space of anther cells of M2BS at anthesis. Moreover, the anther respiration rate and complex I activity of M2BS were significantly lower than those of wild type M2B during pollen development. Furthermore, RNA editing results showed only nad7 presented partially edited at 534th nucleotides. The expression of nad5 and nad7 revealed significant differences between M2B and M2BS. Conclusions Our data demonstrated that mitochondrial structural degradation and complex I deficiency might be associated with transgenic CMS of rice.