Cygb Genes Research Articles

Apoptotic and Oxidative Mechanisms in Liver and Kidney Tissues of Sheep with Fluorosis.

This study was planned to determine the molecular basis and causes of damage to the kidney and the liver, which are the most affected tissues in sheep exposed to chronic fluoride. For this purpose, liver and kidney tissues were obtained from sheep with signs of fluorosis in the age range of 4-6years. The control group consisted of clinically healthy sheep without fluorosis. The apoptotic and oxidative genes expression of target genes was determined using the real qRT-PCR method. According to the control gene (Gapdh) that was detected that in the liver, the apoptotic genes caspase-8, caspase-9, and Bim increased and caspase-3, Bcl-2, and Bak decreased, while in the kidney, caspase-3 and Bax and caspase-8, Bcl-2, Bcl2l-1, Bim, and Bak decreased. According to the 2-ΔCt values of the oxidative stress genes, it was determined that Cygb, Gstp1, and Ncf1 genes increased significantly in the fluorosis group and Gpx1, sod1, and sod2 genes decreased significantly. In the kidney tissue, Cygb and Gpx1 increased in the fluorosis group, while sod1, sod2, Gstp1, Ncf1 and Ccs, and Nos2 were found to decrease significantly. As a result, it was shown that apoptosis and oxidative mechanisms are activated in the liver and the kidney tissues of sheep with fluorosis and these parameters have an important role in understanding the molecular basis of tissue damage in fluorosis.

RNA-Seq analysis of gene expression profiling in human retinal vascular endothelial cells after anti- vascular endothecial growth factor treatment

Objective To observe RNA-Seq analysis of gene expression profiling in human retinal vascular endothelial cells after anti-vascular endothecial growth factor (VEGF) treatment. Methods Cultured the retinal vascular endothelial cells in vitro and logarithmic growth phase cells were used for experiments. The cells were divided into VEGF group and VEGF combined with anti-VEGF drugs group. The VEGF group cells were treated with 50 ng/ml VEGF for 72 h to simulate the high VEGF survival conditions of vascular endothelial cells in diabetic retinopathy. VEGF combined with anti-VEGF drug group cells was treated with 50 ng/ml VEGF and 2.5 μg/ml anti-VEGF drugs for 72 h to imitate the microenvironment of cells following the anti-VEGF drugs treatment, and whole transcriptome sequencing approach was applied to the above two groups of cells through RNA-Seq. Now with biological big data obtained as a basis, to analyze the differentially expressed genes (DEGs). And through enrichment analysis to explain the differential functions of DEGs and their signal pathways. Results The gene expression profiles of the two groups of cells were obtained. Through analysis, 328 DEGs were found, including 194 upregulated and 133 downregulated ones. The functions of DEGs were influenced by regulations over molecular biological process, cellular energy metabolism and protein synthesis, etc. Among these genes, SI,PRX and HPGD were related to protein synthesis, BIRCT to cellular apoptosis, and ABLIM1 and CRB2 to retinal development, and ABCG1, ABCA9 and ABCA12 were associated with the cholesterol of macrophage and the transfer of phospholipid. GO enrichment analysis showed that DEGs mainly act in three ways: regulating biological behavior, organizing cellular component and performing molecular function. Pathway enrichment analysis showed that gene expressions of the two cell groups were differentiated in ECM receptor pathway, and Notch, mitogen-activated protein kinase, transforming growth factor (TGF)-β and Wnt signal pathways. Among them, the gene expression in TGF-β signal pathway attracts most attention, where the DEGs, such as CAMK2B, COL3A1, CYGB, PTGER2 and HS6ST2, among others, were closely related to fibrosis process. Conclusion The anti-VEGF drugs may enhance the expression of CAMK2B, COL3A1, CYGB, PTGER2 and others genes related to TGF-β signal pathway and aggravate retinal fibrosis disease. Key words: Retinal Vessels/cytology; Endothelial cells; Gene expression profiling; Angiogenesis Inhibitors/therapeutic use

1 - Interactions of zebrafish Cytoglobins with oxygen and nitric oxide

Cytoglobin (Cygb) is a ubiquitously expressed heme protein that has been identified in the genome of most higher vertebrate clades including fish, birds and mammals. Since its discovery, extensive study has revealed many structural and functional characteristics of the human Cygb protein, although its physiologic functions remain unknown. Fish have two Cygb genes (Cygb1 and Cygb2); these proteins show small but significant differences, with Cygb2 being more similar to mammalian Cygbs. To gain insights about Cygb function, we compared the properties of zebrafish cytoglobins to the mammalian proteins. Human Cygb exhibits high-affinity, cooperative oxygen binding (P50 of roughly 1 torr, Hill coefficient up to 1.6), as well as nitric oxide (NO) scavenging through NO dioxygenation and NO production via nitrite reduction. These reactions may modulate NO signaling in vivo. Reduced cytochrome b5 (CYB5) has been shown to rapidly reduce Cygb. We have reported that human CYB5 efficiently reduces both human and zebrafish Cygbs, suggesting a conserved role for the CYB5 reduction pathway. In this work, we characterize the two zebrafish Cygbs, measuring key parameters and comparing these results to human Cygb. Specifically, we examine oxygen binding for affinity and cooperativity, NO dioxygenation activity, and the ability of zebrafish CYB5 and CYB5R to reduce zebrafish Cygbs during NO dioxygenation. Oxygen affinity measurements suggest high oxygen affinity, with P50 values of

Molecular characterization, phylogenetic analysis and expression profiling of myoglobin and cytoglobin genes in response to heat stress in channel catfish Ictalurus punctatus.

To understand the function of myoglobin (Mb) and cytoglobin (Cygb) in channel catfish Ictalurus punctatus in response to heat stress, mb and cygb genes were identified and characterized in this study. These genes were widely expressed in all the tested tissues, but strong tissue preferences were observed, with the mb gene being expressed most highly in the heart, cygb1 most highly expressed in the intestine and cygb2 most highly expressed in the brain. After heat-stress challenge, mb and cygb genes were up-regulated in almost all tested tissues. In general, such up-regulation was more dramatic in the tolerant group than in the intolerant group, suggesting that higher expression of mb and cygb genes contributed to greater tolerance of I. punctatus to heat stress.

Differential Loss and Retention of Cytoglobin, Myoglobin, and Globin-E during the Radiation of Vertebrates

If rates of postduplication gene retention are positively correlated with levels of functional constraint, then gene duplicates that have been retained in a restricted number of taxonomic lineages would be expected to exhibit relatively low levels of sequence conservation. Paradoxical patterns are presented by gene duplicates that have been retained in a small number of taxa but which are nonetheless subject to strong purifying selection relative to paralogous members of the same multigene family. This pattern suggests that such genes may have been co-opted for novel, lineage-specific functions. One possible example involves the enigmatic globin-E gene (GbE), which appears to be exclusively restricted to birds. Available data indicate that this gene is expressed exclusively in the avian eye, but its physiological function remains a mystery. In contrast to the highly restricted phyletic distribution of GbE, the overwhelming majority of jawed vertebrates (gnathostomes) possess copies of the related cytoglobin (Cygb) and myoglobin (Mb) genes. The purpose of the present study was 1) to assess the phyletic distribution of the Cygb, Mb, and GbE genes among vertebrates, 2) to elucidate the duplicative origins and evolutionary histories of these three genes, and 3) to evaluate the relative levels of functional constraint of these genes based on comparative sequence analysis. To accomplish these objectives, we conducted a combined phylogenetic and comparative genomic analysis involving taxa that represent each of the major lineages of gnathostome vertebrates. Results of synteny comparisons and phylogenetic topology tests revealed that GbE is clearly not the product of a recent, bird-specific duplication event. Instead, GbE originated via duplication of a proto-Mb gene in the stem lineage of gnathostomes. Unlike the Mb gene, which has been retained in all major gnathostome lineages other than amphibians, the GbE gene has been retained only in the lineage leading to modern birds and has been independently lost in at least four major lineages: teleost fish, amphibians, mammals, and nonavian reptiles. Despite the restricted phyletic distribution of this gene, our results indicate that GbE is one of the most highly conserved globins in the avian genome.

Effect of paraquat exposure on nitric oxide-responsive genes in rat mesencephalic cells

When neural cells are exposed to paraquat, nitric oxide generation increases primarily due to an increase in the expression of the inducible isoform of nitric oxide synthase. The nitric oxide generated has controversial actions in paraquat exposure, as both protective and harmful effects have been described previously. While the actions mediated by nitric oxide in neural cells have been well described, there is evidence that nitric oxide may also be an important modulator of the expression of several genes during paraquat exposure. To better understand the actions of nitric oxide and its potential role in paraquat-induced gene expression, we examined changes in GCH1, ARG1, ARG2, NOS1, NOS2, NOS3, NOSTRIN, NOSIP, NOS1AP, RASD1, DYNLL1, GUCY1A3, DDAH1, DDAH2 and CYGB genes whose expression is controlled by or involved in signaling by the second messenger nitric oxide, in rat mesencephalic cells after 3, 6, 12 and 24 h of paraquat exposure. A qPCR strategy targeting these genes was developed using a SYBR green I-based method. The mRNA levels of all the genes studied were differentially regulated during exposure. These results demonstrate that nitric oxide-related genes are regulated following paraquat exposure of mesencephalic cells and provide the basis for further studies exploring the physiological and functional significance of nitric oxide-sensitive genes in paraquat-mediated neurotoxicity.

Cytoglobin, the Newest Member of the Globin Family, Functions as a Tumor Suppressor Gene

Cytoglobin (CYGB) is a recently discovered vertebrate globin distantly related to myoglobin with unknown function. CYGB is assigned to chromosomal region 17q25, which is frequently lost in multiple malignancies. Previous studies failed to detect evidence for mutations in the CYGB gene. Recent studies provided preliminary evidence for increased methylation of the gene in lung cancer. Our study was aimed at investigating the role of CYGB as a tumor suppressor gene. By nested methylation-specific DNA sequencing analysis of lung and breast cancer cell lines and bronchial and mammary epithelial cell lines, we identified that methylation of a 110-bp CpG-rich segment of the CYGB promoter was correlated with gene silencing. We specifically targeted this sequence and developed a quantitative methylation-specific PCR assay, suitable for high-throughput analysis. We showed that the tumor specificity of CYGB methylation in discriminating patients with and without lung cancer, using biopsies and sputum samples. We further showed the tumor specificity of this assay with multiple other epithelial and hematologic malignancies. To show tumor suppressor activity of CYGB, we performed the following: (a) RNA interference-mediated knockdown of CYGB gene on colony formation in a CYGB expression-positive lung cancer cell line, resulting in increased colony formation; (b) enforced gene expression in CYGB expression-negative lung and breast cancer cell lines, reducing colony formation; and (c) identification of potential proximate targets down-stream of the CYGB genes. Our data constitute the first direct functional evidence for CYGB, the newest member of the globin family, as a tumor suppressor gene.

Quantitative methylation analysis of resection margins and lymph nodes in oral squamous cell carcinoma

Resection is the preferred treatment for oral squamous cell carcinoma, and pathological staging of the resected specimen is crucial. The role of molecular biology in the diagnosis of minimal residual disease has not been fully investigated and may improve staging. Multiple adjacent specimens were taken from the tumour, the invasive front, the surgical margin, and the lymph nodes of 20 specimens from patients with oral cancer. Bisulphite-treated DNA from these specimens was assayed quantitatively with pyrosequencing methylation assays (PMA) of CpG islands within the gene promoters of the p16 and CYGB genes. Results were recorded with histopathological results, and compared with clinical outcome. Biological and technical replicates confirmed the reliability of the techniques. PMA upgraded 13 of the 20 surgical margins, 6 of which subsequently had a recurrent tumour. Not all of these recurrences were predicted and the effects of adjuvant treatment make firm conclusions difficult.

Gene expression and tissue distribution of cytoglobin and myoglobin in the Amphibia and Reptilia: Possible compensation of myoglobin with cytoglobin in skeletal muscle cells of anurans that lack the myoglobin gene

Cytoglobin (Cygb), a recently discovered vertebrate cytoplasmic heme-binding globin, is considered to be in a clade with vertebrate myoglobin (Mb), which is exclusively distributed in the cytoplasm of cardiac and skeletal muscles as an oxygen storage protein. GenBank databases (NCBI and JGI) and gene synteny analyses showed the absence of the Mb gene ( mb) in two anuran amphibians, Xenopus laevis and X. tropicalis. Here we conducted comparative studies on the gene expression and tissue distribution of Cygb and Mb in anuran and reptilian tissues. Cygb and Mb genes were cloned from a reptile, iguana ( Iguana iguana). Two types of cygb ( cygb-1 and -2) were cloned, with lengths of 1066 and 1034 bp, and 196 and 193 amino acid residues, respectively. Their nucleotide and amino acid sequence identities were 90 and 87%, respectively. The Mb gene covered 1416 bp with an open reading frame of 465 bp, giving rise to a 154 amino acid protein. The distal ligand-binding histidine at E7, the proximal heme-binding histidine at F8, and the phenylalanine residue at CD1 were conserved in Mb and Cygb. The nucleotide and amino acid sequence identity of I. iguana cygb-1 and cygb-2 against X. laevis cygb were approximately 67% and 65%, respectively. RT-PCR demonstrated that X. laevis cygb was uniquely expressed in the heart and skeletal muscles, and faintly in the liver and spleen, which was quite contrasted with Iguana and the other vertebrates, where mb is exclusively expressed in the heart and skeletal muscles. Immunohistochemical analyses showed the distribution of Cygb in the cytoplasm of skeletal muscle cells. Interestingly, Cygb in the heart was localized in the nuclei. Considering the absence of mb in the Anura, we hypothesize that Cygb in muscle cells of anurans compensates for the lack of Mb for the storage and intracellular transportation of oxygen.