Biohythane Production in Hydrogen-Oriented Dark Fermentation of Aerobic Granular Sludge (AGS) Pretreated with Solidified Carbon Dioxide (SCO2).

Though deemed a prospective method, the bioconversion of organic waste to biohydrogen via dark fermentation (DF) has multiple drawbacks and limitations. Technological difficulties of hydrogen fermentation may, in part, be eliminated by making DF a viable method for biohythane production. Aerobic granular sludge (AGS) is a little-known organic waste spurring a growing interest in the municipal sector; its characteristics indicate the feasibility of its use as a substrate for biohydrogen production. The major goal of the present study was to determine the effect of AGS pretreatment with solidified carbon dioxide (SCO2) on the yield of H2 (biohythane) production during anaerobic digestion (AD). It was found that an increasing dose of SCO2 caused an increase in concentrations of COD, N-NH4+, and P-PO43- in the supernatant at the SCO2/AGS volume ratios from 0 to 0.3. The AGS pretreatment at SCO2/AGS ratios within the range of 0.1-0.3 was shown to enable the production of biogas with over 8% H2 (biohythane) content. The highest yield of biohythane production, reaching 481 ± 23 cm3/gVS, was obtained at the SCO2/AGS ratio of 0.3. This variant produced 79.0 ± 6% CH4 and 8.9 ± 2% H2. The higher SCO2 doses applied caused a significant decrease in the pH value of AGS, modifying the anaerobic bacterial community to the extent that diminished anaerobic digestion performance.

Identification of a novel homozygous synthesis of cytochrome c oxidase 2 variant in siblings with early-onset axonal Charcot-Marie-Tooth disease.

The synthesis of cytochrome c oxidase 2 (SCO2 ) gene encodes for a mitochondrial located metallochaperone essential for the synthesis of the cytochrome c oxidase (COX) subunit 2. Recessive mutations in SCO2 have been reported in several cases with fatal infantile cardioencephalomyopathy with COX deficiency and in only four cases with axonal neuropathy. Here, we identified a homozygous pathogenic variant (c.361G > C; p.[Gly121Arg]) in SCO2 in two brothers with isolated axonal motor neuropathy. To address pathogenicity of the amino acid substitution, biochemical studies were performed and revealed increased level of the mutant SCO2 -protein and dysregulation of COX subunits in leukocytes and moreover unraveled decrease of proteins involved in the manifestation of neuropathies. Hence, our combined data strengthen the concept of SCO2 being causative for a very rare form of axonal neuropathy, expand its molecular genetic spectrum and provide first biochemical insights into the underlying pathophysiology.

Development and Analysis of the Novel Hybridization of a Single-Flash Geothermal Power Plant with Biomass Driven sCO2-Steam Rankine Combined Cycle.

This study investigates the hybridization scenario of a single-flash geothermal power plant with a biomass-driven sCO2-steam Rankine combined cycle, where a solid local biomass source, olive residue, is used as a fuel. The hybrid power plant is modeled using the simulation software EBSILON®Professional. A topping sCO2 cycle is chosen due to its potential for flexible electricity generation. A synergy between the topping sCO2 and bottoming steam Rankine cycles is achieved by a good temperature match between the coupling heat exchanger, where the waste heat from the topping cycle is utilized in the bottoming cycle. The high-temperature heat addition problem, common in sCO2 cycles, is also eliminated by utilizing the heat in the flue gas in the bottoming cycle. Combined cycle thermal efficiency and a biomass-to-electricity conversion efficiency of 24.9% and 22.4% are achieved, respectively. The corresponding fuel consumption of the hybridized plant is found to be 2.2 kg/s.

Bibliometric Analysis on Supercritical CO2 Power Cycles for Concentrating Solar Power Applications.

In recent years, supercritical CO2 power cycles have received a large amount of interest due to their exceptional theoretical conversion efficiency above 50%, which is leading a revolution in power cycle research. Furthermore, this high efficiency can be achieved at a moderate temperature level, thus suiting concentrating solar power (CSP) applications, which are seen as a core business within supercritical technologies. In this context, numerous studies have been published, creating the need for a thorough analysis to identify research areas of interest and the main researchers in the field. In this work, a bibliometric analysis of supercritical CO2 for CSP applications was undertaken considering all indexed publications within the Web of Science between 1990 and 2020. The main researchers and areas of interest were identified through network mapping and text mining techniques, thus providing the reader with an unbiased overview of sCO2 research activities. The results of the review were compared with the most recent research projects and programs on sCO2 for CSP applications. It was found that popular research areas in this topic are related to optimization and thermodynamics analysis, which reflects the significance of power cycle configuration and working conditions. Growing interest in medium temperature applications and the design of sCO2 heat exchangers was also identified through density visualization maps and confirmed by a review of research projects.

Cotyledons contribute to plant growth and hybrid vigor in Arabidopsis.

MAIN CONCLUSION: In hybrids of Arabidopsis, cotyledons influence the amount and proportion of hybrid vigor in total plant growth. We found Arabidopsis cotyledons are essential for plant growth and in some hybrids for hybrid vigor. In hybrids between C24 and Landsberg erecta (Ler), biomass vigor (heterosis) occurs in the first few days after sowing (DAS), with hybrid cotyledons being larger than those of their parents. C24xLer hybrids are ahead of their parents in activating photosynthesis and auxin pathway genes in cotyledons at 3-4 DAS. "Earliness" is also present in newly emerged C24xLer hybrid leaves. We showed cotyledon removal at 4 DAS caused significant biomass reduction in later growth in hybrids and parental lines. The biomass decrease caused by cotyledon removal can be partially rescued by exogenous sucrose or auxin with different genotypes responding to sucrose and/or auxin differently. Cotyledon removal has different effects on heterosis in different hybrids. After cotyledon removal, in C24xLer hybrids, both growth and heterosis were reduced in similar proportions, but the level of hybrid vigor was reduced as a proportion of growth in C24xColumbia (Col) and ColxLer hybrids. The removal of cotyledons at 4 DAS markedly decreased the level of growth and eliminated the heterotic phenotype of Wassilewskija (Ws)/Ler hybrids. In mutant Ws/Ler hybrids which had a reduced level of photosynthesis in the cotyledons, there was a reduction in plant growth and loss of heterosis. The variation in contribution of cotyledons to heterosis in different hybrids indicates there are multiple pathways to achieve heterotic phenotypes.

Investigating the cardiac pathology of SCO2-mediated hypertrophic cardiomyopathy using patients induced pluripotent stem cell-derived cardiomyocytes.

Mutations in SCO2 are among the most common causes of COX deficiency, resulting in reduced mitochondrial oxidative ATP production capacity, often leading to hypertrophic cardiomyopathy (HCM). To date, none of the recent pertaining reports provide deep understanding of the SCO2 disease pathophysiology. To investigate the cardiac pathology of the disease, we were the first to generate induced pluripotent stem cell (iPSC)-derived cardiomyocytes (iPSC-CMs) from SCO2-mutated patients. For iPSC generation, we reprogrammed skin fibroblasts from two SCO2 patients and healthy controls. The first patient was a compound heterozygote to the common E140K mutation, and the second was homozygote for the less common G193S mutation. iPSC were differentiated into cardiomyocytes through embryoid body (EB) formation. To test the hypothesis that the SCO2 mutation is associated with mitochondrial abnormalities, and intracellular Ca2+ -overload resulting in functional derangements and arrhythmias, we investigated in SCO2-mutated iPSC-CMs (compared to control cardiomyocytes): (i) the ultrastructural changes; (ii) the inotropic responsiveness to ß-adrenergic stimulation, increased [Ca2+ ]o and angiotensin-II (AT-II); and (iii) the Beat Rate Variability (BRV) characteristics. In support of the hypothesis, we found in the mutated iPSC-CMs major ultrastructural abnormalities and markedly attenuated response to the inotropic interventions and caffeine, as well as delayed afterdepolarizations (DADs) and increased BRV, suggesting impaired SR Ca2+ handling due to attenuated SERCA activity caused by ATP shortage. Our novel results show that iPSC-CMs are useful for investigating the pathophysiological mechanisms underlying the SCO2 mutation syndrome.

Human mitochondrial cytochrome c oxidase assembly factor COX18 acts transiently as a membrane insertase within the subunit 2 maturation module.

Defects in mitochondrial cytochrome c oxidase or respiratory chain complex IV (CIV) assembly are a frequent cause of human mitochondrial disorders. Specifically, mutations in four conserved assembly factors impinging the biogenesis of the mitochondrion-encoded catalytic core subunit 2 (COX2) result in myopathies. These factors afford stability of newly synthesized COX2 (the dystonia-ataxia syndrome protein COX20), a protein with two transmembrane domains, and maturation of its copper center, CuA (cardiomyopathy proteins SCO1, SCO2, and COA6). COX18 is an additional COX2 assembly factor that belongs to the Oxa1 family of membrane protein insertases. Here, we used a gene-editing approach to generate a human COX18 knock-out HEK293T cell line that displays isolated complete CIV deficiency. We demonstrate that COX20 stabilizes COX2 during insertion of its N-proximal transmembrane domain, and subsequently, COX18 transiently interacts with COX2 to promote translocation across the inner membrane of the COX2 C-tail that contains the apo-CuA site. The release of COX18 from this complex coincides with the binding of the SCO1-SCO2-COA6 copper metallation module to COX2-COX20 to finalize COX2 biogenesis. Therefore, COX18 is a new candidate when screening for mitochondrial disorders associated with isolated CIV deficiency.

Imatinib inhibits the expression of SCO2 and FRATAXIN genes that encode mitochondrial proteins in human Bcr-Abl⁺ leukemia cells.

Imatinib mesylate (IM/Gleevec®), a selective inhibitor of chimeric Bcr-Abl tyrosine kinase, was developed as a first line drug to treat CML and ALL Ph(+) patients. Earlier studies have shown that hemin counteracts the IM-induced cell killing in human K-562 CML cells. In this study, we investigated whether IM disrupts the heme-dependent Cytochrome c Oxidase (COX) Biosynthesis and Assembly Pathway (HDCBAP) in Bcr-Abl(+) and Bcr-Abl(-) cells by affecting the expression of key-genes. Cells were exposed to IM and evaluated at time intervals for cell growth, cell death, expression of various genes by RT-PCR analysis as well as Sco2 mature protein levels by western blot analysis and COX enzymatic activity. IM at 1 µM induced extensive cell growth inhibition and cell death as well as marked suppression of the expression of SCO2 and FRATAXIN (FXN) genes in human K-562 and KU-812 Bcr-Abl(+) CML cells. IM also reduced the protein level of mature Sco2 mitochondrial protein as well as COX activity in these cell lines. However, treatment of human MOLT-4 Bcr-Abl(-) cells with 1µM and even with higher concentrations (4×10(-5)M) of IM neither reduced the expression of SCO2 and FXN genes nor suppressed the protein level of mature Sco2 protein and COX activity. Our findings indicate that SCO2 and FXN genes, involved in HDCBAP, are repressed by IM in human Bcr-Abl(+) CML cells and may represent novel target sites in leukemia therapy.

Protein Transduction Domain-Mediated Delivery of Recombinant Proteins and In Vitro Transcribed mRNAs for Protein Replacement Therapy of Human Severe Genetic Mitochondrial Disorders: The Case of Sco2 Deficiency.

Mitochondrial disorders represent a heterogeneous group of genetic disorders with variations in severity and clinical outcomes, mostly characterized by respiratory chain dysfunction and abnormal mitochondrial function. More specifically, mutations in the human SCO2 gene, encoding the mitochondrial inner membrane Sco2 cytochrome c oxidase (COX) assembly protein, have been implicated in the mitochondrial disorder fatal infantile cardioencephalomyopathy with COX deficiency. Since an effective treatment is still missing, a protein replacement therapy (PRT) was explored using protein transduction domain (PTD) technology. Therefore, the human recombinant full-length mitochondrial protein Sco2, fused to TAT peptide (a common PTD), was produced (fusion Sco2 protein) and successfully transduced into fibroblasts derived from a SCO2/COX-deficient patient. This PRT contributed to effective COX assembly and partial recovery of COX activity. In mice, radiolabeled fusion Sco2 protein was biodistributed in the peripheral tissues of mice and successfully delivered into their mitochondria. Complementary to that, an mRNA-based therapeutic approach has been more recently considered as an innovative treatment option. In particular, a patented, novel PTD-mediated IVT-mRNA delivery platform was developed and applied in recent research efforts. PTD-IVT-mRNA of full-length SCO2 was successfully transduced into the fibroblasts derived from a SCO2/COX-deficient patient, translated in host ribosomes into a nascent chain of human Sco2, imported into mitochondria, and processed to the mature protein. Consequently, the recovery of reduced COX activity was achieved, thus suggesting the potential of this mRNA-based technology for clinical translation as a PRT for metabolic/genetic disorders. In this review, such research efforts will be comprehensibly presented and discussed to elaborate their potential in clinical application and therapeutic usefulness.

Technological, Ecological, and Energy-Economic Aspects of Using Solidified Carbon Dioxide for Aerobic Granular Sludge Pre-Treatment Prior to Anaerobic Digestion.

The technology of aerobic granular sludge (AGS) seems prospective in wastewater bio-treatment. The characteristics as well as compactness and structure of AGS have been proved to significantly affect the effectiveness of thus far deployed methods for sewage sludge processing, including anaerobic digestion (AD). Therefore, it is deemed necessary to extend knowledge on the possibilities of efficient AGS management and to seek viable technological solutions for methane fermentation of sludge of this type, including by means of using the pre-treatment step. Little is known about the pre-treatment method with solidified carbon dioxide (SCO2), which can be recovered in processes of biogas upgrading and enrichment, leading to biomethane production. This study aimed to determine the impact of AGS pre-treatment with SCO2 on the efficiency of its AD. An energy balance and a simplified economic analysis of the process were also carried out. It was found that an increasing dose of SCO2 applied in the pre-treatment increased the concentrations of COD, N-NH4+, and P-PO43- in the supernatant in the range of the SCO2/AGS volume ratios from 0.0 to 0.3. No statistically significant differences were noted above the latter value. The highest unit yields of biogas and methane production, reaching 476 ± 20 cm3/gVS and 341 ± 13 cm3/gVS, respectively, were obtained in the variant with the SCO2/AGS ratio of 0.3. This experimental variant also produced the highest positive net energy gain, reaching 1047.85 ± 20 kWh/ton total solids (TS). The use of the higher than 0.3 SCO2 doses was proved to significantly reduce the pH of AGS (below 6.5), thereby directly diminishing the percentage of methanogenic bacteria in the anaerobic bacterial community, which in turn contributed to a reduced CH4 fraction in the biogas.

Mitochondrial complex IV defects induce metabolic and signaling perturbations that expose potential vulnerabilities in HCT116 cells.

Mutations in genes encoding cytochrome c oxidase (mitochondrial complex IV) subunits and assembly factors [e.g., synthesis of cytochrome c oxidase 2 (SCO2)] are linked to severe metabolic syndromes. Notwithstanding that SCO2 is under transcriptional control of tumor suppressor p53, the role of mitochondrial complex IV dysfunction in cancer metabolism remains obscure. Herein, we demonstrate that the loss of SCO2 in HCT116 colorectal cancer cells leads to significant metabolic and signaling perturbations. Specifically, abrogation of SCO2 increased NAD+ regenerating reactions and decreased glucose oxidation through citric acid cycle while enhancing pyruvate carboxylation. This was accompanied by a reduction in amino acid levels and the accumulation of lipid droplets. In addition, SCO2 loss resulted in hyperactivation of the insulin-like growth factor 1 receptor (IGF1R)/AKT axis with paradoxical downregulation of mTOR signaling, which was accompanied by increased AMP-activated kinase activity. Accordingly, abrogation of SCO2 expression appears to increase the sensitivity of cells to IGF1R and AKT, but not mTOR inhibitors. Finally, the loss of SCO2 was associated with reduced proliferation and enhanced migration of HCT116 cells. Collectively, herein we describe potential adaptive signaling and metabolic perturbations triggered by mitochondrial complex IV dysfunction.

Development of a novel PTD-mediated IVT-mRNA delivery platform for potential protein replacement therapy of metabolic/genetic disorders.

The potential clinical applications of the powerful in vitro-transcribed (IVT)-mRNAs, to restore defective protein functions, strongly depend on their successful intracellular delivery and transient translation through the development of safe and efficient delivery platforms. In this study, an innovative (international patent-pending) methodology was developed, combining the IVT-mRNAs with the protein transduction domain (PTD) technology, as an efficient delivery platform. Based on the PTD technology, which enables the intracellular delivery of various cargoes intracellularly, successful conjugation of a PTD to the IVT-mRNAs was achieved and evaluated by band-shift assay and NMR spectroscopy. In addition, the PTD-IVT-mRNAs were applied and evaluated in two protein-disease models, including the mitochondrial disorder fatal infantile cardioencephalomyopathy and cytochrome c oxidase (COX) deficiency (attributed to SCO2 gene mutations) and ß-thalassemia. The PTD-IVT-mRNA of SCO2 was successfully transduced and translated to the corresponding Sco2 protein inside the primary fibroblasts of a SCO2/COX-deficient patient, whereas the PTD-IVT-mRNA of ß-globin was transduced and translated in bone marrow cells, derived from three ß-thalassemic patients. The transducibility and the structural stability of the PDT-IVT-mRNAs, in both cases, were confirmed at the RNA and protein levels. We propose that our novel delivery platform could be clinically applicable as a protein therapy for metabolic/genetic disorders.

In vivo biodistribution study of TAT-L-Sco2 fusion protein, developed as protein therapeutic for mitochondrial disorders attributed to SCO2 mutations.

The rapid progress achieved in the development of many biopharmaceuticals had a tremendous impact on the therapy of many metabolic/genetic disorders. This type of fruitful approach, called protein replacement therapy (PRT), aimed to either replace the deficient or malfunctional protein in human tissues that act either in plasma membrane or via a specific cell surface receptor. However, there are also many metabolic/genetic disorders attributed to either deficient or malfunctional proteins acting intracellularly. The recent developments of Protein Transduction Domain (PTD) technology offer new opportunities by allowing the intracellular delivery of recombinant proteins of a given therapeutic interest into different subcellular sites and organelles, such as mitochondria and other entities. Towards this pathway, we applied successfully PTD Technology as a protein therapeutic approach, in vitro, in SCO2 deficient primary fibroblasts, derived from patient with mutations in human SCO2 gene, responsible for fatal, infantile cardioencephalomyopathy and cytochrome c oxidase deficiency. In this work, we radiolabeled the recombinant TAT-L-Sco2 fusion protein with technetium-99 m to assess its in vivo biodistribution and fate, by increasing the sensitivity of detection of even low levels of the transduced recombinant protein. The biodistribution pattern of [99mTc]Tc-TAT-L-Sco2 in mice demonstrated fast blood clearance, significant hepatobiliary and renal clearance. In addition, western blot analysis detected the recombinant TAT-L-Sco2 protein in the isolated mitochondria of several mouse tissues, including heart, muscle and brain. These results pave the way to further consider this PTD-mediated Protein Therapy Approach as a potentially alternative treatment of genetic/metabolic disorders.

COA6 Is Structurally Tuned to Function as a Thiol-Disulfide Oxidoreductase in Copper Delivery to Mitochondrial Cytochrome c Oxidase.

In eukaryotes, cellular respiration is driven by mitochondrial cytochrome c oxidase (CcO), an enzyme complex that requires copper cofactors for its catalytic activity. Insertion of copper into its catalytically active subunits, including COX2, is a complex process that requires metallochaperones and redox proteins including SCO1, SCO2, and COA6, a recently discovered protein whose molecular function is unknown. To uncover the molecular mechanism by which COA6 and SCO proteins mediate copper delivery to COX2, we have solved the solution structure of COA6, which reveals a coiled-coil-helix-coiled-coil-helix domain typical of redox-active proteins found in the mitochondrial inter-membrane space. Accordingly, we demonstrate that COA6 can reduce the copper-coordinating disulfides of its client proteins, SCO1 and COX2, allowing for copper binding. Finally, our determination of the interaction surfaces and reduction potentials of COA6 and its client proteins provides a mechanism of how metallochaperone and disulfide reductase activities are coordinated to deliver copper to CcO.

High expression of synthesis of cytochrome c oxidase 2 and TP53-induced glycolysis and apoptosis regulator can predict poor prognosis in human lung adenocarcinoma.

Synthesis of cytochrome c oxidase 2 (SCO2) and TP53-induced glycolysis and apoptosis regulator (TIGAR) are 2 p53-mediated proteins that can play a regulatory role in cancer energy metabolism. However, no study has examined the association of SCO2 and TIGAR with the prognosis of patients with lung adenocarcinoma (AC). In our study, the expression of SCO2 and TIGAR proteins in lung AC was detected, and the potential relation to prognosis was evaluated, aiming to take a further view of lung AC progression. Quantum dots-based immunofluorescence histochemistry staining was performed to observe the expression of p53, SCO2, and TIGAR in 75 specimens of lung AC. Of these, 51 (68.0%) showed high expression of SCO2, and 59 (78.7%) showed high expression of TIGAR. High TIGAR expression was significantly associated with a history of smoking (P = .017) and being male (P = .006). The correlation between high SCO2 expression and age also was significant (P = .042). Moreover, high TIGAR expression was positively correlated with high SCO2 expression (P = .019; rs = 0.271). High expression of the SCO2 and TIGAR proteins predicted poorer survival and a higher mortality rate (P = .024 and .030, respectively). High expression of SCO2 and TIGAR proteins is significantly associated with lung AC progression, suggesting their potential use as prognostic markers and therapeutic targets.

Sco2 deficient mice develop increased adiposity and insulin resistance.

Cytochrome c oxidase (COX) is an essential transmembrane protein complex (Complex IV) in the mitochondrial respiratory electron chain. Mutations in genes responsible for the assembly of COX are associated with Leigh syndrome, cardiomyopathy, spinal muscular atrophy and other fatal metabolic disorders in humans. Previous studies have shown that mice lacking the COX assembly protein Surf1 (Surf1-/- mice) paradoxically show a number of beneficial metabolic phenotypes including increased insulin sensitivity, upregulation of mitochondrial biogenesis, induction of stress response pathways and increased lifespan. To determine whether these effects are specific to the Surf1 mutation or a more general effect of reduced COX activity, we asked whether a different mutation causing reduced COX activity would have similar molecular and physiologic changes. Sco2 knock-in/knock-out (KI/KO) mice in which one allele of the Sco2 gene that encodes a copper chaperone required for COX activity is deleted and the second allele is mutated, have previously been shown to be viable despite a 30-60% reduction in COX activity. In contrast to the Surf1-/- mice, we show that Sco2 KI/KO mice have increased fat mass, associated with reduced ß-oxidation and increased adipogenesis markers, reduced insulin receptor beta (IR-ß levels in adipose tissue, reduced muscle glucose transporter 4 (Glut4) levels and a impaired response to the insulin tolerance test consistent with insulin resistance. COX activity and protein are reduced approximately 50% in adipose tissue from the Sco2 KI/KO mice. Consistent with the increase in adipose tissue mass, the Sco2 KI/KO mice also show increased hepatosteatosis, elevated serum and liver triglyceride and increased serum cholesterol levels compared to wild-type controls. In contrast to the Surf1-/- mice, which show increased mitochondrial number, upregulation of the mitochondrial unfolded protein response (UPRMT) pathway and no significant change in mitochondrial respiration in several tissues, Sco2 KI/KO mice do not upregulate the UPRMT, and tissue oxygen consumption and levels of several proteins involved in mitochondrial function are reduced in adipose tissue compared to wild type mice. Thus, the metabolic effects of the Sco2 and Surf1-/- mutations are opposite, despite comparable changes in COX activity, illuminating the complex impact of mitochondrial dysfunction on physiology and pointing to an important role for complex IV in regulating metabolism.

SNOWY COTYLEDON 2 Promotes Chloroplast Development and Has a Role in Leaf Variegation in Both Lotus japonicus and Arabidopsis thaliana.

Plants contain various factors that transiently interact with subunits or intermediates of the thylakoid multiprotein complexes, promoting their stable association and integration. Hence, assembly factors are essential for chloroplast development and the transition from heterotrophic to phototrophic growth. Snowy cotyledon 2 (SCO2) is a DNAJ-like protein involved in thylakoid membrane biogenesis and interacts with the light-harvesting chlorophyll-binding protein LHCB1. In Arabidopsis thaliana, SCO2 function was previously reported to be restricted to cotyledons. Here we show that disruption of SCO2 in Lotus japonicus results not only in paler cotyledons but also in variegated true leaves. Furthermore, smaller and pale-green true leaves can also be observed in A. thaliana sco2 (atsco2) mutants under short-day conditions. In both species, SCO2 is required for proper accumulation of PSII-LHCII complexes. In contrast to other variegated mutants, inhibition of chloroplastic translation strongly affects L. japonicus sco2 mutant development and fails to suppress their variegated phenotype. Moreover, inactivation of the suppressor of variegation AtClpR1 in the atsco2 background results in an additive double-mutant phenotype with variegated true leaves. Taken together, our results indicate that SCO2 plays a distinct role in PSII assembly or repair and constitutes a novel factor involved in leaf variegation.

Low energy costs of F1Fo ATP synthase reversal in colon carcinoma cells deficient in mitochondrial complex IV.

Mitochondrial polarisation is paramount for a variety of cellular functions. Under ischemia, mitochondrial membrane potential (ΔΨm) and proton gradient (ΔpH) are maintained via a reversal of mitochondrial F1Fo ATP synthase (mATPase), which can rapidly deplete ATP and drive cells into energy crisis. We found that under normal conditions in cells with disassembled cytochrome c oxidase complex (COX-deficient HCT116), mATPase maintains ΔΨm at levels only 15-20% lower than in WT cells, and for this utilises relatively little ATP. For a small energy expenditure, mATPase enables mitochondrial ΔpH, protein import, Ca2+ turnover, and supports free radical detoxication machinery enlarged to protect the cells from oxidative damage. Whereas in COX-deficient cells the main source of ATP is glycolysis, the ΔΨm is still maintained upon inhibition of the adenine nucleotide translocators with bongkrekic acid and carboxyatractyloside, indicating that the role of ANTs is redundant, and matrix substrate level phosphorylation alone or in cooperation with ATP-Mg/Pi carriers can continuously support the mATPase activity. Intriguingly, we found that mitochondrial complex III is active, and it contributes not only to free radical production, but also to ΔΨm maintenance and energy budget of COX-deficient cells. Overall, this study demonstrates that F1Fo ATP synthase can support general mitochondrial and cellular functions, working in extremely efficient 'energy saving' reverse mode and flexibly recruiting free radical detoxication and ATP producing / transporting pathways.

Association between SCO2 mutation and extreme myopia in Japanese patients.

PURPOSE: To investigate the role of SCO2 in extreme myopia of Japanese patients. METHODS: In total, 101 Japanese patients with extreme myopia (axial length of ≥30 mm) OU at the Kyoto University Hospital were included in this study. Exon 2 of SCO2 was sequenced by conventional Sanger sequencing. The detected variants were assessed using in silico prediction programs: SIFT, PolyPhen-2 and MutationTaster. To determine the frequency of the mutations in normal subjects, we referred to the 1000 Genomes Project data and the Human Genetic Variation Database (HGVD) in the Human Genetic Variation Browser. RESULTS: The average age of the participants was 62.9 ± 12.7 years. There were 31 males (30.7 %) and 70 females. Axial lengths were 31.76 ± 1.17 mm OD and 31.40 ± 1.07 mm OS, and 176 eyes (87.6 %) out of 201 eyes had myopic maculopathy of grade 2 or more. Among the 101 extremely myopic patients, one mutation (c.290 C > T;p.Ala97Val) in SCO2 was detected. This mutation was not found in the 1000 Genomes Project data or HGVD data. Variant type of the mutation was nonsynonymous. Although the SIFT prediction score was 0.350, the PolyPhen-2 probability was 0.846, thus predicting its pathogenicity to be possibly damaging. MutationTaster PhyloP was 1.268, suggesting that the mutation is conserved. CONCLUSIONS: We identified one novel possibility of an extreme myopia-causing mutation in SCO2. No other disease-causing mutation was found in 101 extremely myopic Japanese patients, suggesting that SCO2 plays a limited role in Japanese extreme myopia. Further investigation is required for better understanding of extreme myopia.

The African-specific S47 polymorphism of p53 alters chemosensitivity.

The TP53 protein is known to affect the sensitivity of tumor cells to cell death by DNA damaging agents. We recently reported that human and mouse cells containing an African-specific coding region variant of p53, Pro47Ser (hereafter S47), are impaired in the transactivation of a small subset of p53 target genes including GLS2 and SCO2, and are markedly resistant to cisplatin. Further, mice containing this variant are markedly predisposed to cancer. Together these findings suggested that cancer-affected humans with the S47 variant might not be effectively treated with cisplatin. To more directly test this premise, we created transformed derivatives of mouse embryo fibroblasts (MEFs) containing wild type p53 (WT) and the S47 variant and analyzed them for chemosensitivity. We find that transformation with E1A and Ras actually reverses the chemosensitivity/transcriptional differences between WT p53 and S47. Specifically, E1A/Ras-transformed S47 cells show increased sensitivity to cisplatin and paclitaxel, and comparable transactivation of GLS2 and SCO2, compared to cells with WT p53. These data suggest that the functional differences between WT p53 and S47 in primary cells may not hold true for transformed cells. They also offer hope that cisplatin and paclitaxel may be effective chemotherapeutic drugs for S47 individuals with cancer.