Oxidase Subunit VIc Research Articles

Potential evidence for epigenetic biomarkers of metabolic syndrome in human whole blood in Latinos.

Metabolic syndrome (MetS) is highly prevalent worldwide. In the United States, estimates show that more than 30% of the adult population has MetS. MetS consists of multiple phenotypes, including obesity, dyslipidemia, and impaired glucose tolerance. Therefore, identifying the molecular mechanisms to explain this complex disease is critical for diagnosing and treating MetS. We previously showed 70 increased genes and 20 decreased genes in whole blood in MetS participants. The present study aimed to identify blood-based DNA methylation biomarkers in non-MetS versus MetS participants. The present study analyzed whole blood DNA samples from 184 adult participants of Latino descent from the Arizona Insulin Resistance (AIR) registry. We used the National Cholesterol Education Program Adult Treatment Panel III (NCEP: ATP III) criteria to identify non-MetS (n = 110) and MetS (n = 74) participants. We performed whole blood methylation analysis on select genes: ATP Synthase, H+ Transporting mitochondrial F1 Complex, Epsilon Subunit (ATP5E), Cytochrome C Oxidase Subunit VIc (COX6C), and Ribosomal Protein L9 (RPL9). The pyrosequencing analysis was a targeted approach focusing on the promoter region of each gene that specifically captured CpG methylation sites. In MetS participants, we showed decreased methylation in two CpG sites in COX6C and three CpG sites in RPL9, all p < 0.05 using the Mann-Whitney U test. There were no ATP5E CpG sites differently methylated in the MetS participants. Furthermore, while adjusting for age, gender, and smoking status, logistic regression analysis reaffirmed the associations between MetS and mean methylation within COX6C and RPL9 (both p < 0.05). In addition, Spearman’s correlation revealed a significant inverse relationship between the previously published gene expression data and methylation data for RPL9 (p < 0.05). In summary, these results highlight potential blood DNA methylation biomarkers for the MetS phenotype. However, future validation studies are warranted to strengthen our findings.

Estrogen Enhances the Expression of the Multidrug Transporter Gene ABCG2-Increasing Drug Resistance of Breast Cancer Cells through Estrogen Receptors.

Background: Multidrug resistance is a major obstacle in the successful therapy of breast cancer. Studies have proved that this kind of drug resistance happens in both human cancers and cultured cancer cell lines. Understanding the molecular mechanisms of drug resistance is important for the reasonable design and use of new treatment strategies to effectively confront cancers. Results: In our study, ATP-binding cassette sub-family G member 2 (ABCG2), adenosine triphosphate (ATP) synthase and cytochrome c oxidase subunit VIc (COX6C) were over-expressed more in the MCF-7/MX cell line than in the normal MCF7 cell line. Therefore, we believe that these three genes increase the tolerance of MCF7 to mitoxantrone (MX). The data showed that the high expression of COX6C made MCF-7/MX have more stable on mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) expression than normal MCF7 cells under hypoxic conditions. The accumulation of MX was greater in the ATP-depleted treatment MCF7/MX cells than in normal MCF7/MX cells. Furthermore, E2 increased the tolerance of MCF7 cells to MX through inducing the expression of ABCG2. However, E2 could not increase the expression of ABCG2 after the inhibition of estrogen receptor α (ERα) in MCF7 cells. According to the above data, under the E2 treatment, MDA-MB231, which lacks ER, had a higher sensitivity to MX than MCF7 cells. Conclusions: E2 induced the expression of ABCG2 through ERα and the over-expressed ABCG2 made MCF7 more tolerant to MX. Moreover, the over-expressed ATP synthase and COX6c affected mitochondrial genes and function causing the over-expressed ABCG2 cells pumped out MX in a concentration gradient from the cell matrix. Finally lead to chemoresistance.

Effects of moderate global maternal nutrient reduction on fetal baboon renal mitochondrial gene expression at 0.9 gestation.

Early life malnutrition results in structural alterations in the kidney, predisposing offspring to later life renal dysfunction. Kidneys of adults who were growth restricted at birth have substantial variations in nephron endowment. Animal models have indicated renal structural and functional consequences in offspring exposed to suboptimal intrauterine nutrition. Mitochondrial bioenergetics play a key role in renal energy metabolism, growth, and function. We hypothesized that moderate maternal nutrient reduction (MNR) would adversely impact fetal renal mitochondrial expression in a well-established nonhuman primate model that produces intrauterine growth reduction at term. Female baboons were fed normal chow diet or 70% of control diet (MNR). Fetal kidneys were harvested at cesarean section at 0.9 gestation (165 days gestation). Human Mitochondrial Energy Metabolism and Human Mitochondria Pathway PCR Arrays were used to analyze mitochondrially relevant mRNA expression. In situ protein content was detected by immunohistochemistry. Despite the smaller overall size, the fetal kidney weight-to-body weight ratio was not affected. We demonstrated fetal sex-specific differential mRNA expression encoding mitochondrial metabolite transport and dynamics proteins. MNR-related differential gene expression was more evident in female fetuses, with 16 transcripts significantly altered, including 14 downregulated and 2 upregulated transcripts. MNR impacted 10 transcripts in male fetuses, with 7 downregulated and 3 upregulated transcripts. The alteration in mRNA levels was accompanied by a decrease in mitochondrial protein cytochrome c oxidase subunit VIc. In conclusion, transcripts encoding fetal renal mitochondrial energy metabolism proteins are nutrition sensitive in a sex-dependent manner. We speculate that these differences lead to decreased mitochondrial fitness that contributes to renal dysfunction in later life.

Abstract 292: Isoform-Specific Expression of Estrogen-Related Receptors Regulates Cardiomyocyte Metabolic Adaptation to Oxidative Stress

In the ischemic heart, metabolic adaptation of cardiomyocytes exposed to prolonged hypoxia is critical to sustain myocardial energy production. Metabolic gene expression is regulated by Estrogen-Related Receptor (ERR) transcription factors and Peroxisome Proliferator-Activated Receptor γ Coactivator-1 (PGC-1) co-activators. ERRs do not require ligand to achieve an active conformation and regulate an array of target genes involved in myocardial energy metabolism. These studies aim to determine the regulatory mechanisms governing expression of ERR isoforms (ERRα and ERRβ) in cardiomyocytes undergoing oxidative stress. We hypothesize that metabolic energy production in myocardium initially adapts to hypoxia by inducing transient ERRβ expression, and maintains this compensatory expression through induction of sustained ERRα expression. Adult feline cardiomyocytes in primary culture were electrically stimulated to contract continuously at 1 Hz under normoxia (21% [O2]), hypoxia (0.5% [O2]), or hypoxia followed by reoxygenation. Expression of ERR, PGC-1 and target genes was measured by QRT-PCR (n≥5). QRT-PCR arrays were used to screen for prospective target genes. In response to 24 hours of hypoxia, mRNA levels increased for ERRα (3-fold by 24 hours), ERRβ (11-fold by 12 hours), PGC-1α (16-fold by 24 hours) and PGC-1β (8-fold by 24 hours). Subsequent reoxygenation for 24 hours decreased ERRβ, PGC-1α and PGC-1β mRNA levels to that of normoxia controls. However, reoxygenation did not markedly decrease (∼25%; 0.7 of 3.0-fold) ERRα mRNA levels. Cytochrome C Oxidase Subunit VIc (Cox6c) and Fatty Acid Binding Protein 3 (Fabp3) were identified as potential ERRα target genes in adult cardiomyocytes based on correlation of expression. Moreover, hypoxia-induced increases in Cox6c and Fabp3 mRNA levels were partially blocked by treatment with XCT-790, an inhibitor of ERRα activity. We conclude that hypoxia and subsequent reoxygenation elicit isoform-specific changes in ERR and PGC-1 mRNA expression in adult cardiomyocytes. This observed regulation of expression for ERR isoforms and their target genes may play a critical role in metabolic adaptation to oxidative stress.

Gene expression profiling in glomeruli of diabetic nephropathy rat

Diabetic nephropathy (DN) remains the most common cause of end-stage renal disease (ESRD) as the burden of diabetes increases worldwide. To find improved intervention strategies for this disease, it is necessary to investigate the molecular mechanisms involved. To obtain more insight into processes that lead to DN, mRNA expression profiles of diabetic and normal glomeruli from rat kidneys were compared. Rats were divided into a control group and a DN group randomly. The DN group was injected with streptozotocin. Fasting blood glucose (FBG) and weight were measured monthly. On the 12th week, blood samples were collected and analyzed for plasma creatinine and blood urea nitrogen (BUN). Glomeruli were isolated and Illumina Rat Ref-12 V1.0 Expression Beadchip gene array was performed. Quantitative realtime polymerase chain reaction (Q-RT-PCR) was used to confirm the results of gene array for a selected number of genes. We found FBG, 24-h urinary albumin, serum creatinine and BUN were significantly increased, while urinary creatinine and body weight were significantly decreased in the DN group. Glomeruli from the DN group had 624 genes with differential expression. DAVID (Database for Annotation, Visualization and integrated Discovery) analysis showed that the three most enriched terms were 'cytosol' (GO:0005829), 'translational elongation' (GO:0006414) and 'mitochondion' (GO:0005739). Those genes could be mapped to eight pathways. The most common type of enriched pathway was related to 'extracellular matrix (ECM)-receptor interaction'. Other pathways included those for 'ribosome', 'focal adhesion', 'oxidative phosphorylation', 'transforming growth factor (TGF)-beta signaling pathway', 'Parkinson's disease', 'Alzheimer's disease' and 'renin-angiotensin system'. Q-RT-PCR verified that Atp5b (F1-ATPase beta subunit), Col1a1 (collagen type 1 alpha 1), Cox6c (cytochrome c oxidase subunit VIc), Ndufs3 (NADH dehydrogenase [ubiquinone] Fe-S protein 3) and Tgfb1 (transforming growth factor β1) were significantly up-regulated in the DN group. The expressions of NDUFS3 and TGF-β1 in DN rats were increased. Our findings suggested that the oxidative phosphorylation pathway, ECM-receptor interaction, TGF-β pathway and renin-angiotensin system may be involved in the development of DN.

Telmisartan improves kidney function through inhibition of the oxidative phosphorylation pathway in diabetic rats

Telmisartan provides renal benefit at all stages of the renal continuum in patients with type 2 diabetes mellitus. This research is to investigate the effect of telmisartan on kidney function in diabetic rats and to identify the underlying molecular mechanisms. Diabetic rats were divided into vehicle group, low dosage (TeL) group, and high dosage of telmisartan (TeH) group. We performed Illumina RatRef-12 Expression BeadChip gene array experiments. We found 3-months of treatment with telmisartan significantly decreased 24-h urinary albumin, serum creatinine, blood urea nitrogen, and increased creatinine clearance rate. Kidney hypertrophy and glomerular mesangial matrix expansion were ameliorated. The glomeruli from the TeH group had 1541 genes with significantly changed expression (554 increased, 987 decreased). DAVID (Database for annotation, visualization and Integrated discovery) analyses showed that the most enriched term was 'mitochondrion' (Gene Ontology (GO:0005739)) in all 67 GO functional categories. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses indicated that all differentially expressed genes included seven KEGG pathways. Of those pathways, four are closely related to the oxidative phosphorylation pathway. Quantitative real-time PCR verified that the H+ transporting mitochondrial F1 complex, beta subunit (Atp5b), cytochrome c oxidase subunit VIc (Cox6c), and NADH dehydrogenase (ubiquinone) Fe-S protein 3 (Ndufs3) were significantly downregulated both in TeL and TeH groups, while nephrosis 1 homolog (Nphs1) and nephrosis 2 homolog (Nphs2) were significantly upregulated. The increased expression of malonaldehyde and NDUFS3 in the glomeruli of diabetic rats was attenuated by telmisartan. The other significantly changed pathway we found was the peroxisome proliferator-activated receptor (PPAR) signaling pathway. Our data suggest that telmisartan can improve kidney function in diabetic rats. The mechanism may be involved in mitochondrion oxidative phosphorylation, the PPAR-γ pathway, and the slit diaphragm.

COX6C (cytochrome c oxidase subunit VIc)

Review on COX6C (cytochrome c oxidase subunit VIc), with data on DNA, on the protein encoded, and where the gene is implicated.

Identification of differentially expressed genes in senescence-accelerated mouse testes by suppression subtractive hybridization analysis

Senescence-accelerated mouse (SAM) strains constitute a model of accelerated senescence coupled with a short lifespan and the early development of various age-related disorders. To identify differential gene expression in testes between senescence-accelerated SAMP1 and control SAMR1 mice, we performed suppression subtractive hybridization. We observed that the expression of three genes related to cell proliferation (myosin regulatory light chain B, aldolase 1A isoform, and cytochrome c oxidase subunit VIc) were upregulated and four genes implicated in spermatogenesis were downregulated in SAMP1 mice. Asb-8, a member of ankyrin repeat-containing proteins, was abundantly expressed in the testes and downregulated in SAMP1. The other three downregulated genes (germ cell-specific gene 1, T-complex polypeptide 1b, and activator of cAMP responsive element modulator in testis) have been reported to regulate late-stage spermatogenesis. These gene expression profiles might explain the findings of early testicular maturation and rapid decline in the ability to produce spermatozoa with advancing age in SAMP1 mice.

Mass spectrometric identification of mitochondrial oxidative phosphorylation subunits separated by two-dimensional blue-native polyacrylamide gel electrophoresis.

Blue-native polyacrylamide gel electrophoresis is a powerful tool for the separation of intact membrane protein complexes mainly applied to the analysis of the enzymes of the mitochondrial oxidative phosphorylation system (OXPHOS). Combined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), it reveals a two-dimensional pattern showing the individual subunits of the five OXPHOS multi-enzyme complexes. This pattern is useful in the diagnostic analysis of several diseases related to disorders in the oxidative phosphorylation system. However, in order to use this method for systematic diagnostic purposes and to be able to link disease with absence or reduced expression of specific subunits, an unambiguous identification of the individual subunits is necessary. In this study, we completed this task, implementing peptide mass fingerprinting and mass spectrometric sequence analysis. In the course of these analyses, we discovered a novel variant of a cytochrome c oxidase subunit VIc.

The cyclope gene of Drosophila encodes a cytochrome c oxidase subunit VIc homolog.

Cytochrome c oxidase is the terminal enzyme of the mitochondrial electron transfer chain. In eukaryotes, the enzyme is composed of 3 mitochondrial DNA-encoded subunits and 7-10 (in mammals) nuclear DNA-encoded subunits. This enzyme has been extensively studied in mammals and yeast but, in Drosophila, very little is known and no mutant has been described so far. Here we report the genetic and molecular characterization of mutations in cyclope (cype) and the cloning of the gene encoding a cytochrome c oxidase subunit VIc homolog. cype is an essential gene whose mutations are lethal and show pleiotropic phenotypes. The 77-amino acid peptide encoded by cype is 46% identical and 59% similar to the human subunit (75 amino acids). The transcripts are expressed maternally and throughout development in localized regions. They are found predominantly in the central nervous system of the embryo; in the central region of imaginal discs; in the germarium, follicular, and nurse cells of the ovary; and in testis. A search in the Genome Annotation Database of Drosophila revealed the absence of subunit VIIb and the presence of 9 putative nuclear cytochrome c oxidase subunits with high identity scores when compared to the 10 human subunits.

Novel gene fusion of COX6C at 8q22-23 to HMGIC at 12q15 in a uterine leiomyoma.

Cytogenetic analyses have shown that aberrations involving 12q13-15 are frequent chromosomal changes in a variety of human benign mesenchymal tumors, e.g., pleomorphic adenomas of the parotid gland, pulmonary chondroid hamartomas, lipomas, and uterine leiomyomas. Recently, the high-mobility group protein gene HMGIC was identified as the target gene affected by the 12q13-15 aberrations. Using 3' rapid amplification of cDNA ends experiments, we isolated novel ectopic sequences fused to HMGIC in a uterine leiomyoma. Cloning of the fusion cDNA identified the human cytochrome c oxidase subunit VIc (COX6C) gene on 8q22-23 as the fusion partner of HMGIC. Nucleotide sequences of the fusion transcript revealed that the first 3 exons of the HMGIC gene, encoding the 3 DNA binding domains, was fused to the exon 2 of the COX6C gene. The identification of a gene rearrangement suggests a role for HMGIC in tumorigenesis of uterine leiomyoma and suggests a possible involvement of HMGIC in mesenchymal differentiation. Genes Chromosomes Cancer 27:303-307, 2000.

Tissue-Specific Stability of Nuclear- and Mitochondrially Encoded mRNAs

Steady-state levels of mRNAs encoding mitochondrial proteins are drastically different among tissues. We evaluated tissue-specific variations in mRNA stability by comparing rates of mRNA decay in liver, heart, and muscle following the inhibition of transcription. Rates of decline of the mRNAs encoding δ-aminolevulinate synthase (ALAs), cytochrome c oxidase subunit VIc (nuclear-encoded), and subunit III (mitochondrially encoded) in heart, liver, and muscle for 6 h following transcription inhibition with actinomycin D or ethidium bromide were measured. Subunit VIc mRNA levels were least stable in liver (t1/2= 2.4 h), slightly greater in heart (t1/2= 3.3 h), and very stable in skeletal muscle. Similarly, ALAs mRNA exhibited at1/2of 41 min in liver, but this was markedly increased to approximately 11–14 h in heart and skeletal muscle. In contrast, subunit III was least stable in heart (t1/2= 2.1 h), somewhat more stable in liver (t1/2= 3.8 h), but no decline in subunit III mRNA levels occurred in muscle following the inhibition of transcription. Thus, muscle, heart, and liver possess tissue-specific mechanisms which control the stability of mRNAs encoding mitochondrial proteins. In addition, the coordinated expression of subunit III and VIc mRNAs in different tissues is partly due to parallel rates of mRNA turnover. This suggests the presence of intra- and extramitochondrial factors within a tissue which regulate the stability of specific mRNAs in a similar manner.

Nucleotide sequence of cDNA encoding human cytochrome c oxidase subunit VIc.

Nucleotide sequence of cDNA encoding human cytochrome c oxidase subunit VIc.

A processed pseudogene with an intact coding sequence for rat liver cytochrome c oxidase subunit VIc

The biogenesis of eukaryotic cytochrome c oxidase involves the coordinate expression of nuclear and mitochondrial genes. Very little information is available on the gene structure of nuclear-coded cytochrome c oxidase subunits in mammalian systems. We report here the isolation and complete nucleotide sequence determination of a processed pseudogene for cytochrome c oxidase subunit VIc from rat liver. The pseudogene lacks introns and the coding region is intact with no deleterious lesions; however, there are 7 amino acid (aa) differences when compared to the sequence derived from cDNA clones. The pseudogene has the potential to code for a protein of 76 aa, containing a putative 3 aa N-terminal presequence when compared to the mature bovine heart VIc subunit. Potential regulatory regions, including a TATA box, are present in the 5′-flanking region.

Organization and nucleotide sequence of two chromosomal genes for rat cytochrome c oxidase subunit VIc: a structural and a processed gene.

A rat genomic library was screened with a cDNA probe coding for rat liver cytochrome c oxidase subunit VIc (COX-VIc). Out of 16 clones mapped, four inserts were different and did not overlap. Two inserts that gave strong hybridization signals were characterized by sequence analysis. These data show that we have cloned two different genes for rat COX-VIc. The rat COX-VIc-2 gene contains four exons and three introns. The 5' boundary of the first exon was mapped by primer extension experiments. The nucleotide sequence of the first three exons is identical to the sequence of the rat liver cDNA probe; thus, we can conclude that this gene is expressed in rat liver. The second gene (COX-VIc-1) is 88% homologous to the rat liver cDNA and contains an open reading frame of 228 nucleotides capable of coding for a full-length COX-VIc polypeptide of 76 amino acids. This predicted translation product is 79% homologous to the rat liver peptide. The absence of introns and other factors, such as the presence of a 13-bp direct repeat and a poly(A) addition signal, indicate that this locus is a processed gene which may have evolved from spliced mRNA intermediates.