COX7A2L/SCAFI and Pre-Complex III Modify Respiratory Chain Supercomplex Formation in Different Mouse Strains with a Bcs1/Mutation

The COX7A2L (Supercomplex Assembly Factor I, SCAFI) protein has been proposed to be a mitochondrial supercomplex assembly factor required for respirasome (supercomplex containing complexes I, III, and IV) formation. In the C57BU6 mouse strain a homozygous in-frame 6-base-pair deletion in the COX7a2I/SCAF1 gene resulting in unstable protein and suggesting loss of function was previously identified. The loss of SCAFI was shown to impede respirasome formation, a major concern for the use of C57BL mouse strains in mitochondrial research. In contradiction, another recent study suggested that supercomplex formation is independent of SCAFI isoforms. We investigated whether SCAFI isoform status affected the disease severity and supercomplex formation in the liver of Bcs1a232A>G knock-in mice with incomplete complex III assembly. In homozygotes (Bcs1/(G/G)) of mixed (C57BL/6:129/Sv) genetic background, the lifespan was similar in mice with wild-type SCAFI allele and in those homozygous (SCAF/(short/short)) for the deleted SCAF1 variant (34 3 days; n = 6 vs. 32 +/- 2 days; n = 7, respectively). SCAFI heterozygosity (SCAF/(long/short)) resulted in decreased SCAFI protein but respirasome assembly was unaffected. Congenic (C57BL/6) mice were of the genotype SCAF(short/short) and had no detectable SCAFI protein. In their liver mitochondria, respirasome composition was altered as compared to mixed background mice. Complex IV was mainly present as monomers and dimers, and only low amounts were found in combination with complex I and complex III or with precomplex III. The main supercomplex in the liver mitochondria of C57BU6 mice comprised only complexes I and III. In conclusion, in liver mitochondria of C57BL/6 mice, supercomplexes had markedly reduced amount of, but were not completely depleted of, complex IV, supporting a role for COX7A2L/SCAFI in supercomplex assembly. However, the disease progression of the Bcs1/mutant mice was unrelated to SCAFI isoforms and supercomplex composition, suggesting that other genetic factors contribute to the different survival in the different genetic backgrounds.Peer reviewe

Complex I Function and Supercomplex Formation Are Preserved in Liver Mitochondria Despite Progressive Complex III Deficiency

Peer reviewe

The radical-binding lipocalin A1M binds to a Complex I subunit and protects mitochondrial structure and function.

Aims: During cell death, energy-consuming cell degradation and recycling programs are performed. Maintenance of energy-delivery during cell death is therefore crucial but the mechanisms to keep the mitochondrial functions intact during these processes are poorly understood. We have investigated the hypothesis that the heme- and radical-binding ubiquitous protein A1M (α1-microglobulin) is involved in protection of the mitochondria against oxidative insult during cell death. Results: Using blood cells, keratinocytes and liver cells, we show that A1M binds with high affinity to apoptosis-induced cells and is localized to mitochondria. The mitochondrial Complex I subunit NDUFAB1 was identified as a major molecular target of the A1M-binding. Furthermore, A1M was shown to inhibit the swelling of mitochondria, and to reverse the severely abrogated ATP-production of mitochondria when exposed to heme and ROS. Innovation: Import of the radical- and heme-binding protein A1M from the extracellular compartment confers protection of mitochondrial structure and function during cellular insult. Conclusion: A1M binds to a subunit of Complex I and has a role in assisting the mitochondria to maintain its energy delivery during cell death. A1M may also, at the same time, counteract and eliminate the ROS generated by the mitochondrial respiration to prevent oxidative damage to surrounding healthy tissue

Metabolite Profiles Reveal Energy Failure and Impaired Beta-Oxidation in Liver of Mice with Complex III Deficiency Due to a BCS1L Mutation

WOS:000306956300061Peer reviewe

Effect of High-Carbohydrate Diet on Plasma Metabolome in Mice with Mitochondrial Respiratory Chain Complex III Deficiency

Mitochondrial disorders cause energy failure and metabolic derangements. Metabolome profiling in patients and animal models may identify affected metabolic pathways and reveal new biomarkers of disease progression. Using liver metabolomics we have shown a starvation-like condition in a knock-in (Bcs1l(c.232A>G)) mouse model of GRACILE syndrome, a neonatal lethal respiratory chain complex III dysfunction with hepatopathy. Here, we hypothesized that a high-carbohydrate diet (HCD, 60% dextrose) will alleviate the hypoglycemia and promote survival of the sick mice. However, when fed HCD the homozygotes had shorter survival (mean +/- SD, 29 +/- 2.5 days, n = 21) than those on standard diet (33 +/- 3.8 days, n = 30), and no improvement in hypoglycemia or liver glycogen depletion. We investigated the plasma metabolome of the HCD- and control diet-fed mice and found that several amino acids and urea cycle intermediates were increased, and arginine, carnitines, succinate, and purine catabolites decreased in the homozygotes. Despite reduced survival the increase in aromatic amino acids, an indicator of liver mitochondrial dysfunction, was normalized on HCD. Quantitative enrichment analysis revealed that glycine, serine and threonine metabolism, phenylalanine and tyrosine metabolism, and urea cycle were also partly normalized on HCD. This dietary intervention revealed an unexpected adverse effect of high-glucose diet in complex III deficiency, and suggests that plasma metabolomics is a valuable tool in evaluation of therapies in mitochondrial disorders.Peer reviewe

Fasting reveals largely intact systemic lipid mobilization mechanisms in respiratory chain complex III deficient mice

Mice homozygous for the human GRACILE syndrome mutation (Bcs1l (c.A232G)) display decreased respiratory chain complex III activity, liver dysfunction, hypoglycemia, rapid loss of white adipose tissue and early death. To assess the underlying mechanism of the lipodystrophy in homozygous mice (Bcs1l(p.S)(78G)), these and wild-type control mice were subjected to a short 4-hour fast. The homozygotes had low baseline blood glucose values, but a similar decrease in response to fasting as in wild-type mice, resulting in hypoglycemia in the majority. Despite the already depleted glycogen and increased triacylglycerol content in the mutant livers, the mice responded to fasting by further depletion and increase, respectively. Increased plasma free fatty acids (FAs) upon fasting suggested normal capacity for mobilization of lipids from white adipose tissue into circulation. Strikingly, however, serum glycerol concentration was not increased concomitantly with free FM, suggesting its rapid uptake into the liver and utilization for fuel or gluconeogenesis in the mutants. The mutant hepatocyte mitochondria were capable of responding to fasting by appropriate morphological changes, as analyzed by electron microscopy, and by increasing respiration. Mutants showed increased hepatic gene expression of major metabolic controllers typically associated with fasting response (Ppargc1a, Fgf21, Cd36) already in the fed state, suggesting a chronic starvation-like metabolic condition. Despite this, the mutant mice responded largely normally to fasting by increasing hepatic respiration and switching to FA utilization, indicating that the mechanisms driving these adaptations are not compromised by the CIII dysfunction. Summary statement: Bcs1l mutant mice with severe CIII deficiency, energy deprivation and post-weaning lipolysis respond to fasting similarly to wild-type mice, suggesting largely normal systemic lipid mobilization and utilization mechanisms.Peer reviewe

Mechanisms of phagocytosis resistance in Streptococcus pyogenes

A distinguishing feature of Streptococcus pyogenes (GAS) is their ability to resist phagocytosis in whole human blood in the absence of type-specific antibodies, a property that is dependent on the expression of the surface associated M proteins. A common characteristic of M proteins is their interaction with a variety of host proteins including the complement regulatory protein factor H (FH) and fibrinogen that plays a key role in coagulation. Here, we report that GAS also interact with Factor H-like protein-1 (FHL-1), an alternative splice product of FH comprising the first seven short consensus repeats (SCRs) of the FH molecule. Furthermore, we have localised the binding site for the M protein in SCR 7 of FH and FHL-1. Although FH and FHL-1 are very similar, GAS, at a low bacteria to plasma ratio reminiscent of the physiological situation, selectively absorbed FHL-1 but not FH from plasma. Further analysis demonstrated a major binding site for FH/FHL-1 in the A- repeat region of the streptococcal M protein. Unexpectedly, deletion of this region in the M5 protein did not influence the ability of streptococci expressing this protein to resist phagocytosis in human blood. While the interaction with FH/FHL-1 is restricted to a certain subset of GAS strains, the binding of fibrinogen is common to all GAS strains. We mapped the fibrinogen-binding regions in the M1 and M5 proteins to the so-called B-repeats of these proteins. Furthermore, we could demonstrate that these regions contribute to streptococcal survival in human blood thereby suggesting that fibrinogen plays an important role for resistance against phagocytosis Traditionally, resistance against phagocytosis is correlated to the expression of the M proteins. However, we found, that the antiphagocytic property of M proteins is restricted by the genetic background of the strain expressing the particular M protein suggesting that phagocytosis resistance may require cooperation between M proteins and other strain-specific components

Mitochondrial hepatopathies in the newborn period.

Mitochondrial disorders recognized in the neonatal period usually present as a metabolic crisis combined with one or several organ manifestations. Liver disorder in association with a respiratory chain deficiency may be overlooked since liver dysfunction is common in severely sick newborn infants. Lactacidosis, hypoglycemia, elevated serum transaminases and conjugated bilirubin are common signs of mitochondrial hepatopathy. Hepatosplenomegaly may occur in severe cases. A clinical picture with fetal growth restriction, postnatal lactacidosis, hypoglycemia, coagulopathy, and cholestasis, especially in combination with neurological symptoms or renal tubulopathy, should alert the neonatologist to direct investigations on mitochondrial disorder. A normal lactate level does not exclude respiratory chain defects. The most common liver manifestation caused by mutated mitochondrial DNA (deletion) is Pearson syndrome. Recently, mutations in several nuclear DNA genes have been identified that lead to mitochondrial hepatopathy, e.g. mitochondrial depletion syndrome caused by DGUOK, MPV17, SUCLG1, POLG1, or C10ORF2 mutations. A combination of lactacidosis, liver involvement, and Fanconi type renal tubulopathy is common when the complex III assembly factor BCS1L harbors mutations, the most severe disease with consistent genotype-phenotype correlation being the GRACILE syndrome. Mutations in nuclear translation factor genes (TRMU, EFG1, and EFTu) of the respiratory chain enzyme complexes have recently been identified. Diagnostic work-up of neonatal liver disorder should include assessment of function and structure of the complexes as well as mutation screening for known genes. So far, treatment is mainly symptomatic