Maintenance of complex I and its super-complexes by NDUF-11 is essential for mitochondrial structure, function and health

Mitochondrial super-complexes form around a conserved core of monomeric complex I and dimeric complex III; wherein subunit NDUFA11, of the former, is conspicuously situated at the interface. We identified B0491.5 (NDUF-11) as the C. elegans homologue, of which animals homozygous for a CRISPR-Cas9 generated knockout allele arrested at the L2 development stage. Reducing (but not eliminating) expression by RNAi allowed development to adulthood, enabling characterisation of the consequences: destabilisation of complex I and its super-complexes, and perturbation of respiratory function. The loss of NADH-dehydrogenase activity is compensated by enhanced complex II activity, with the potential for detrimental ROS-production. Electron cryo-tomography highlight aberrant cristae morphology and inter-membrane-space widening and cristae-junctions. The requirement of NDUF-11 for balanced respiration, mitochondrial morphology and development presumably arises due to its involvement in complex I/ super-complex maintenance. This highlights the importance of respiratory complex integrity for health and the potential of its perturbation for mitochondrial disease.

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Maintenance of Complex I and respiratory super-complexes by NDUF-11 is essential for respiratory function, mitochondrial structure and health in C. elegans

10.1101/2021.01.06.425530 ◽

2021 ◽

Author(s):

Amber Knapp-Wilson ◽

Gonçalo C. Pereira ◽

Emma Buzzard ◽

Andrew Richardson ◽

Robin A. Corey ◽

...

Keyword(s):

Respiratory Function ◽

Complex I ◽

Metabolic Stress ◽

Development Stage ◽

Complex Iii ◽

Mitochondrial Morphology ◽

C Elegans ◽

Monomeric Complex ◽

L2 Development ◽

Conserved Core

ABSTRACTMitochondrial super-complexes form around a conserved core of monomeric complex I and dimeric complex III; wherein subunit NDUFA11, of the former, is conspicuously situated at the interface. We identified B0491.5 (NDUF-11) as the C. elegans homologue, of which animals homozygous for a CRISPR-Cas9 generated knockout allele arrested at the L2 development stage. Reducing expression by RNAi allowed development to the adult stage, enabling characterisation of the consequences: destabilisation of complex I and its super-complexes, and perturbation of respiratory function. The loss of NADH-dehydrogenase activity is compensated by enhanced complex II activity, resulting in excessive detrimental ROS production. Meanwhile, electron cryo-tomography highlight aberrant cristae morphology and widening of the inter-membrane space and cristae junctions. The requirement of NDUF-11 for balanced respiration, mitochondrial morphology and development highlights the importance of complex I/ super-complex maintenance. Their perturbation by this, or other means, is likely to be the cause of metabolic stress and disease.

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The 12.3 kDa subunit of complex I (respiratory-chain NADH dehydrogenase) from Neurospora crassa: cDNA cloning and chromosomal mapping of the gene

Biochemical Journal ◽

10.1042/bj2910729 ◽

1993 ◽

Vol 291 (3) ◽

pp. 729-732 ◽

Cited By ~ 18

Author(s):

A Videira ◽

J E Azevedo ◽

S Werner ◽

P Cabral

Keyword(s):

Respiratory Chain ◽

Complex I ◽

Nadh Dehydrogenase ◽

Single Gene ◽

Complex Iii ◽

Chromosomal Mapping ◽

Helical Conformation ◽

Significant Similarity ◽

Hydrophobic Domain ◽

Group I

The 12.3 kDa subunit of complex I (respiratory-chain NADH dehydrogenase) is a nuclear-coded protein of the hydrophobic fragment of the enzyme. We have isolated and sequenced a full-length cDNA clone coding for this polypeptide. The deduced protein is 104 amino acid residues long with a molecular mass of 12305 Da. This particular subunit of complex I lacks a cleavable mitochondrial targeting sequence. In agreement with its localization within complex I, we have found that this subunit behaves like an intrinsic membrane protein. Nevertheless, the deduced protein is rather hydrophilic, exhibiting no hydrophobic domain long enough to traverse a membrane in an alpha-helical conformation. The 12.3 kDa subunit shows a significant similarity to the hinge protein of complex III, suggesting that these two polypeptides may be involved in identical functions. This complex I subunit is coded for by a single gene. Applying restriction-fragment-length-polymorphism mapping, we located the gene on the right side of the centromere in linkage group I, linked to the lys-4 locus.

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Effects of proteolytic digestion by chymotrypsin on the structure and catalytic properties of reduced nicotinamide–adenine dinucleotide–ubiquinone oxidoreductase from bovine heart mitochondria

Biochemical Journal ◽

10.1042/bj1650295 ◽

1977 ◽

Vol 165 (2) ◽

pp. 295-301 ◽

Cited By ~ 11

Author(s):

Susan E. Crowder ◽

C. Ian Ragan

Keyword(s):

Complex I ◽

Time Course ◽

Nadh Dehydrogenase ◽

Sodium Dodecyl ◽

Complex Iii ◽

Type Ii ◽

Oxidoreductase Activity ◽

Ubiquinone Oxidoreductase ◽

Nadh Cytochrome ◽

Reduced Nicotinamide Adenine Dinucleotide

1. Incubation of NADH–ubiquinone oxidoreductase (Complex I) with chymotrypsin caused loss of rotenone-sensitive ubiquinone-1 reduction and an increase in rotenone-insensitive ubiquinone reduction. 2. Within the same time-course, NADH–K3Fe(CN)6 oxidoreductase activity was unaffected. 3. Mixing of chymotrypsin-treated Complex I with Complex III did not give rise to NADH–cytochrome c oxidoreductase activity. 4. Gel electrophoresis in the presence of sodium dodecyl sulphate revealed selective degradation of several constituent polypeptides by chymotrypsin. 5. With higher chymotrypsin concentrations and longer incubation times, a decrease in NADH–K3Fe(CN)6 oxidoreductase was observed. The kinetics of this decrease correlated with solubilization of the low-molecular-weight type-II NADH dehydrogenase (subunit mol.wts. 53000 and 27000) and with degradation of a polypeptide of mol.wt. 30000. 6. Phospholipid-depleted Complex I was more rapidly degraded by chymotrypsin. Specifically, a subunit of mol.wt. 75000, resistant to chymotrypsin in untreated Complex I, was degraded in phospholipid-depleted Complex I. In addition, the 30000-mol.wt. polypeptide was also more rapidly digested, correlating with an increased rate of transformation to type II NADH dehydrogenase.

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Beta-carotene Oxygenase 2 (BCO2) Regulates the Mitochondrial Dynamics in Mice (FS06-06-19)

Current Developments in Nutrition ◽

10.1093/cdn/nzz029.fs06-06-19 ◽

2019 ◽

Vol 3 (Supplement_1) ◽

Author(s):

Dingbo Lin ◽

Lei Wu ◽

Xin Guo ◽

Siau Yen Wong ◽

Peiran Lu ◽

...

Keyword(s):

Oxidative Stress ◽

Complex I ◽

Respiratory Activity ◽

Mitochondrial Dynamics ◽

Beta Carotene ◽

Complex Iii ◽

Mitochondrial Morphology ◽

Chow Diet ◽

Wild Type ◽

Mitochondrial Respiratory

Abstract Objectives Beta-carotene oxygenase 2 (BCO2) is a carotenoid metabolic enzyme located to the inner membrane of mitochondria. Decreased expression of and mutations in BCO2 are associated with obesity and metabolic disorders in humans and mice. We recently reported that depletion of BCO2 alters mitochondrial respiratory activity. Here, we further present that BCO2 is important for mitochondrial dynamics and respiratory supercomplex formation in mice. Methods Six-week-old male and female 129S6 (wild type, WT) and BCO2 knockout (KO) mice fed a chow diet were used in the current study. Hypothalamic tissues were collected for mitochondrial morphology by transmission electron microscopy, mitochondrial proteomics, and mitochondrial respiratory supercomplex formation and respiratory activity assays. Key proteins in mitochondrial dynamics, including OPA1, Mfn2, DRP1, p62, ULK1 were assessed by Western blot. Cardiolipin was measured by ELISA. Results The results showed that mitochondrial complex I subunit NDUFA11 was more abundant, assembly of complex I into the I-III-IV-containing supercomplexes was greatly enhanced, yet complex III homodimer was diminished in the hypothalamus of the BCO2 knockout mouse, compared to the wild type. Decreases in mitochondrial respiration activities, disruption of mitochondrial elongation (e.g., increased DRP1 and pS757-ULK1), suppression of mitochondrial biogenesis (e.g., decreased PPARgamma and PGC-1alpha), promotion of mitochondrial oxidative stress, and elevation of the cardiolipin level occurred in depletion of BCO2. Conclusions BCO2 is critical for the hypothalamic mitochondrial homeostasis through regulation of respiratory supercomplex formation, mitochondrial dynamics, and consequent oxidative stress. Funding Sources N/A.

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Effects of lithium treatment on oxidative stress markers in mitochondrial complex I and complex III inhibition and after CO2 exposure in SH-SY5Y cells

10.31487/j.nnb.2019.01.001 ◽

2019 ◽

Author(s):

Frederic Marmol

Keyword(s):

Oxidative Stress ◽

Complex I ◽

Lithium Treatment ◽

Complex Iii ◽

Mitochondrial Complex I ◽

Oxidative Stress Markers ◽

Mitochondrial Complex ◽

Stress Markers

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Neuronal mitochondrial dynamics coordinate systemic mitochondrial morphology and stress response to confer pathogen resistance in C. elegans

Developmental Cell ◽

10.1016/j.devcel.2021.04.021 ◽

2021 ◽

Author(s):

Li-Tzu Chen ◽

Chih-Ta Lin ◽

Liang-Yi Lin ◽

Jiun-Min Hsu ◽

Yu-Chun Wu ◽

...

Keyword(s):

Stress Response ◽

Mitochondrial Dynamics ◽

Pathogen Resistance ◽

Mitochondrial Morphology ◽

C Elegans

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Heterochronic Parabiosis: Old Blood Induces Changes in Mitochondrial Structure and Function of Young Mice

The Journals of Gerontology Series A ◽

10.1093/gerona/glaa299 ◽

2020 ◽

Author(s):

Jenny L Gonzalez-Armenta ◽

Ning Li ◽

Rae-Ling Lee ◽

Baisong Lu ◽

Anthony J A Molina

Keyword(s):

Mitochondrial Respiration ◽

Complex I ◽

Structure And Function ◽

Muscle Performance ◽

Mitochondrial Morphology ◽

Positive Effects ◽

Mitochondrial Structure ◽

And Function ◽

Old Mice ◽

Young Mice

Abstract Heterochronic parabiosis models have been utilized to demonstrate the role of blood-borne circulating factors in systemic effects of aging. In previous studies, heterochronic parabiosis has shown positive effects across multiple tissues in old mice. More recently, a study demonstrated old blood had a more profound negative effect on muscle performance and neurogenesis of young mice. In this study, we used heterochronic parabiosis to test the hypothesis that circulating factors mediate mitochondrial bioenergetic decline, a well-established biological hallmark of aging. We examined mitochondrial morphology, expression of mitochondrial complexes, and mitochondrial respiration from skeletal muscle of mice connected as heterochronic pairs, as well as young and old isochronic controls. Our results indicate that young heterochronic mice had significantly lower total mitochondrial content and on average had significantly smaller mitochondria compared to young isochronic controls. Expression of complex IV followed a similar pattern: young heterochronic mice had a trend for lower expression compared to young isochronic controls. Additionally, respirometric analyses indicate that young heterochronic mice had significantly lower complex I, complex I + II, and maximal mitochondrial respiration and a trend for lower complex II-driven respiration compared to young isochronic controls. Interestingly, we did not observe significant improvements in old heterochronic mice compared to old isochronic controls, demonstrating the profound deleterious effects of circulating factors from old mice on mitochondrial structure and function. We also found no significant differences between the young and old heterochronic mice, demonstrating that circulating factors can be a driver of age-related differences in mitochondrial structure and function.

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

International Journal of Molecular Sciences ◽

10.3390/ijms22126524 ◽

2021 ◽

Vol 22 (12) ◽

pp. 6524

Author(s):

Bo-Yu Lin ◽

Gui-Teng Zheng ◽

Kai-Wen Teng ◽

Juan-Yu Chang ◽

Chao-Chang Lee ◽

...

Keyword(s):

Membrane Potential ◽

Fusion Proteins ◽

Complex I ◽

Nadh Dehydrogenase ◽

Leigh Syndrome ◽

Protein Transduction ◽

Independent Manner ◽

Complex I Deficiency ◽

Transactivator Of Transcription ◽

Hiv 1

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.

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