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1.
Electron. j. biotechnol ; 41: 37-47, sept. 2019. tab, graf, ilus
Article in English | LILACS | ID: biblio-1087161

ABSTRACT

Background: Circular RNAs, a novel class in the eukaryotic transcriptome, are characterized by the 3' and 5' ends that are covalently joined in a covalently closed loop without free ends. Circular RNAs are considerably stable molecules and act as microRNA sponges with regulatory potential to the protein-coding genes. Results: Eight circular RNAs were found to be significantly upregulated at anagen skin tissue of cashmere goat compared with their counterparts at telogen. Rich and complex regulatory patterns were revealed among the eight upregulated circular RNAs at anagen and related miRNAs with their potential regulatory genes. The potential regulatory genes of eight upregulated circular RNAs at anagen were involved in several pathways related to the main physiological process of hair follicle, such as histone acetylation and axon. For chi_circ_1926, chi_circ_3541, chi_circ_0483, chi_circ_3196, and chi_circ_2092, overall, the relative expression in secondary hair follicle exhibited highly similar trends with their corresponding host genes during the different stages of the hair follicle cycle. However, the expression trends of chi_circ_0100, chi_circ_2829, and chi_circ_1967 were found to diverge from their corresponding host genes during the different stages of the hair follicle cycle. Conclusions: A total of eighteen circular RNAs were identified and characterized from skin tissue of cashmere goat. The eight upregulated circular RNAs at anagen might have significant roles in the secondary hair follicle of cashmere goat. Our results would provide a novel regulatory layer to elucidate the molecular mechanisms underlying the development of secondary hair follicle and the growth of cashmere fiber in cashmere goat.


Subject(s)
Animals , Goats/genetics , Hair Follicle/growth & development , RNA, Circular/genetics , Skin , Gene Expression , Computational Biology , MicroRNAs , Eukaryotic Cells , Gene Regulatory Networks , Transcriptome , RNA, Circular/metabolism
2.
Article in English | WPRIM | ID: wpr-719481

ABSTRACT

The Endoplasmic reticulum (ER), an indispensable sub-cellular component of the eukaryotic cell carries out essential functions, is critical to the survival of the organism. The chaperone proteins and the folding enzymes which are multi-domain ER effectors carry out 3-dimensional conformation of nascent polypeptides and check misfolded protein aggregation, easing the exit of functional proteins from the ER. Diverse conditions, for instance redox imbalance, alterations in ionic calcium levels, and inflammatory signaling can perturb the functioning of the ER, leading to a build-up of unfolded or misfolded proteins in the lumen. This results in ER stress, and aiming to reinstate protein homeostasis, a well conserved reaction called the unfolded protein response (UPR) is elicited. Equally, in protracted cellular stress or inadequate compensatory reaction, UPR pathway leads to cell loss. Dysfunctional ER mechanisms are responsible for neuronal degeneration in numerous human diseases, for instance Alzheimer's, Parkinson's and Huntington's diseases. In addition, mounting proof indicates that ER stress is incriminated in psychiatric diseases like major depressive disorder, bipolar disorder, and schizophrenia. Accumulating evidence suggests that pharmacological agents regulating the working of ER may have a role in diminishing advancing neuronal dysfunction in neuropsychiatric disorders. Here, new findings are examined which link the foremost mechanisms connecting ER stress and cell homeostasis. Furthermore, a supposed new pathogenic model of major neuropsychiatry disorders is provided, with ER stress proposed as the pivotal step in disease development.


Subject(s)
Apoptosis , Biological Psychiatry , Bipolar Disorder , Calcium , Depressive Disorder, Major , Endoplasmic Reticulum Stress , Endoplasmic Reticulum , Eukaryotic Cells , Homeostasis , Humans , Neurons , Neuropsychiatry , Oxidation-Reduction , Peptides , Proteostasis Deficiencies , Schizophrenia , Unfolded Protein Response
3.
Article in English | WPRIM | ID: wpr-741766

ABSTRACT

OBJECTIVES: The emergence of resistant bacteria is being increasingly reported around the world, potentially threatening millions of lives. Amongst resistant bacteria, methicillin-resistant Staphylococcus aureus (MRSA) is the most challenging to treat. This is due to emergent MRSA strains and less effective traditional antibiotic therapies to Staphylococcal infections. The use of bacteriophages (phages) against MRSA is a new, potential alternate therapy. In this study, morphology, genetic and protein structure of lytic phages against MRSA have been analysed. METHODS: Isolation of livestock and sewage bacteriophages were performed using 0.4 μm membrane filters. Plaque assays were used to determine phage quantification by double layer agar method. Pure plaques were then amplified for further characterization. Sulfate-polyacrylamide gel electrophoresis and random amplification of polymorphic DNA were run for protein evaluation, and genotyping respectively. Transmission electron microscope was also used to detect the structure and taxonomic classification of phage visually. RESULTS: Head and tail morphology of bacteriophages against MRSA were identified by transmission electron microscopy and assigned to the Siphoviridae family and the Caudovirales order. CONCLUSION: Bacteriophages are the most abundant microorganism on Earth and coexist with the bacterial population. They can destroy bacterial cells successfully and effectively. They cannot enter mammalian cells which saves the eukaryotic cells from lytic phage activity. In conclusion, phage therapy may have many potential applications in microbiology and human medicine with no side effect on eukaryotic cells.


Subject(s)
Agar , Bacteria , Bacteriophages , Caudovirales , Classification , DNA , Electrophoresis , Eukaryotic Cells , Head , Humans , Livestock , Membranes , Methicillin Resistance , Methicillin-Resistant Staphylococcus aureus , Methods , Microscopy, Electron, Scanning Transmission , Microscopy, Electron, Transmission , Sewage , Siphoviridae , Staphylococcal Infections , Tail
4.
Article in English | WPRIM | ID: wpr-765353

ABSTRACT

The mechanistic target of rapamycin (mTOR) pathway coordinates the metabolic activity of eukaryotic cells through environmental signals, including nutrients, energy, growth factors, and oxygen. In the nervous system, the mTOR pathway regulates fundamental biological processes associated with neural development and neurodegeneration. Intriguingly, genes that constitute the mTOR pathway have been found to be germline and somatic mutation from patients with various epileptic disorders. Hyperactivation of the mTOR pathway due to said mutations has garnered increasing attention as culprits of these conditions : somatic mutations, in particular, in epileptic foci have recently been identified as a major genetic cause of intractable focal epilepsy, such as focal cortical dysplasia. Meanwhile, epilepsy models with aberrant activation of the mTOR pathway have helped elucidate the role of the mTOR pathway in epileptogenesis, and evidence from epilepsy models of human mutations recapitulating the features of epileptic patients has indicated that mTOR inhibitors may be of use in treating epilepsy associated with mutations in mTOR pathway genes. Here, we review recent advances in the molecular and genetic understanding of mTOR signaling in epileptic disorders. In particular, we focus on the development of and limitations to therapies targeting the mTOR pathway to treat epileptic seizures. We also discuss future perspectives on mTOR inhibition therapies and special diagnostic methods for intractable epilepsies caused by brain somatic mutations.


Subject(s)
Biological Phenomena , Brain , Drug Resistant Epilepsy , Epilepsies, Partial , Epilepsy , Eukaryotic Cells , Humans , Intercellular Signaling Peptides and Proteins , Malformations of Cortical Development , Nervous System , Oxygen , Sirolimus
5.
Article in English | WPRIM | ID: wpr-788781

ABSTRACT

The mechanistic target of rapamycin (mTOR) pathway coordinates the metabolic activity of eukaryotic cells through environmental signals, including nutrients, energy, growth factors, and oxygen. In the nervous system, the mTOR pathway regulates fundamental biological processes associated with neural development and neurodegeneration. Intriguingly, genes that constitute the mTOR pathway have been found to be germline and somatic mutation from patients with various epileptic disorders. Hyperactivation of the mTOR pathway due to said mutations has garnered increasing attention as culprits of these conditions : somatic mutations, in particular, in epileptic foci have recently been identified as a major genetic cause of intractable focal epilepsy, such as focal cortical dysplasia. Meanwhile, epilepsy models with aberrant activation of the mTOR pathway have helped elucidate the role of the mTOR pathway in epileptogenesis, and evidence from epilepsy models of human mutations recapitulating the features of epileptic patients has indicated that mTOR inhibitors may be of use in treating epilepsy associated with mutations in mTOR pathway genes. Here, we review recent advances in the molecular and genetic understanding of mTOR signaling in epileptic disorders. In particular, we focus on the development of and limitations to therapies targeting the mTOR pathway to treat epileptic seizures. We also discuss future perspectives on mTOR inhibition therapies and special diagnostic methods for intractable epilepsies caused by brain somatic mutations.


Subject(s)
Biological Phenomena , Brain , Drug Resistant Epilepsy , Epilepsies, Partial , Epilepsy , Eukaryotic Cells , Humans , Intercellular Signaling Peptides and Proteins , Malformations of Cortical Development , Nervous System , Oxygen , Sirolimus
6.
Article in English | WPRIM | ID: wpr-129010

ABSTRACT

Mitochondria play essential role in eukaryotic cells including in the oxidative phosphorylation and generation of adenosine triphosphate via the electron-transport chain. Therefore, defects in mitochondrial DNA (mtDNA) can result in mitochondrial dysfunction which leads to various mitochondrial disorders that may present with various neurologic and non-neurologic manifestations. Mutations in the nuclear gene polymerase gamma (POLG) are associated with mtDNA depletions, and Alpers-Huttenlocher syndrome is one of the most severe manifestations of POLG mutation characterized by the clinical triad of intractable seizures, psychomotor regression, and liver failure. The hepatic manifestation usually occurs late in the disease's course, but in some references, hepatitis was reportedly the first manifestation. Liver transplantation was considered contraindicated in Alpers-Huttenlocher syndrome due to its poor prognosis. We acknowledged a patient with the first manifestation of the disease being hepatic failure who eventually underwent liver transplantation, and whose neurological outcome improved after cocktail therapy.


Subject(s)
Adenosine Triphosphate , Diffuse Cerebral Sclerosis of Schilder , DNA, Mitochondrial , Eukaryotic Cells , Hepatitis , Humans , Liver Failure , Liver Transplantation , Liver , Mitochondria , Mitochondrial Diseases , Oxidative Phosphorylation , Prognosis , Seizures
7.
Article in English | WPRIM | ID: wpr-128994

ABSTRACT

Mitochondria play essential role in eukaryotic cells including in the oxidative phosphorylation and generation of adenosine triphosphate via the electron-transport chain. Therefore, defects in mitochondrial DNA (mtDNA) can result in mitochondrial dysfunction which leads to various mitochondrial disorders that may present with various neurologic and non-neurologic manifestations. Mutations in the nuclear gene polymerase gamma (POLG) are associated with mtDNA depletions, and Alpers-Huttenlocher syndrome is one of the most severe manifestations of POLG mutation characterized by the clinical triad of intractable seizures, psychomotor regression, and liver failure. The hepatic manifestation usually occurs late in the disease's course, but in some references, hepatitis was reportedly the first manifestation. Liver transplantation was considered contraindicated in Alpers-Huttenlocher syndrome due to its poor prognosis. We acknowledged a patient with the first manifestation of the disease being hepatic failure who eventually underwent liver transplantation, and whose neurological outcome improved after cocktail therapy.


Subject(s)
Adenosine Triphosphate , Diffuse Cerebral Sclerosis of Schilder , DNA, Mitochondrial , Eukaryotic Cells , Hepatitis , Humans , Liver Failure , Liver Transplantation , Liver , Mitochondria , Mitochondrial Diseases , Oxidative Phosphorylation , Prognosis , Seizures
8.
Article in English | WPRIM | ID: wpr-50094

ABSTRACT

Protein arginine methyltransferase (PRMT) is an important epigenetic regulator in eukaryotic cells. During encystation, an essential process for Acanthamoeba survival, the expression of a lot of genes involved in the encystation process has to be regulated in order to be induced or inhibited. However, the regulation mechanism of these genes is yet unknown. In this study, the full-length 1,059 bp cDNA sequence of Acanthamoeba castellanii PRMT1 (AcPRMT1) was cloned for the first time. The AcPRMT1 protein comprised of 352 amino acids with a SAM-dependent methyltransferase PRMT-type domain. The expression level of AcPRMT1 was highly increased during encystation of A. castellanii. The EGFP-AcPRMT1 fusion protein was distributed over the cytoplasm, but it was mainly localized in the nucleus of Acanthamoeba. Knock down of AcPRMT1 by synthetic siRNA with a complementary sequence failed to form mature cysts. These findings suggested that AcPRMT1 plays a critical role in the regulation of encystation of A. castellanii. The target gene of AcPRMT1 regulation and the detailed mechanisms need to be investigated by further studies.


Subject(s)
Acanthamoeba castellanii , Acanthamoeba , Amino Acids , Clone Cells , Cytoplasm , DNA, Complementary , Epigenomics , Eukaryotic Cells , Protein-Arginine N-Methyltransferases , RNA, Small Interfering
9.
Article in Chinese | WPRIM | ID: wpr-309112

ABSTRACT

<p><b>OBJECTIVE</b>This study aimed to construct a eukaryotic expression vector pEGFP-N1-EMP1 of epithelial mem-brane protein 1 (EMP1) and investigate its influence on migration and invasion of human oral tongue squamous carcinoma cells.</p><p><b>METHODS</b>The human EMP1 gene was amplified by reverse transcription polymerase chain reaction and then ligated into the pEGFP-N1 vector by double restriction endonuclease digestion to construct pEGFP-N1-EMP1 recombinant plasmid. After sequencing identification, pEGFP-N1-EMP1 recombinant plasmid and pEGFP-N1 plasmid were transfected into human oral tongue squamous carcinoma Tb3.1 cell line. The expression of green fluorescent protein in cells was observed after transfection using an inverted fluorescence microscope. The overexpression of EMP1 mRNA was identified at 24, 48, and 72 h after transfection by real-time fluorescence quantitative polymerase chain reaction. The effect of EMP1 overexpression on migration and invasion of Tb3.1 cells was detected by Transwell assay.</p><p><b>RESULTS</b>The full-length EMP1 gene sequence was successfully obtained. Sequence analysis showed that the EMP1 gene was inserted into the pEGFP-N1 vector correctly. Green fluorescence was observed in the transfected cells under fluorescence microscopy. The results of real-time fluorescence quantitative polymerase chain reaction indicated that the expression of EMP1 at 24 h after pEGFP-N1-EMP1 transfection was significantly higher than the other groups. Transwell assays indicated that overexpression of the EMP1 gene could significantly inhibit the migration and invasion ability of Tb3.1 cells.</p><p><b>CONCLUSIONS</b>The eukaryotic expression vector of EMP1 was successfully constructed, and EMP1 overexpression was confirmed to inhibit the migration and inva-sion of oral tongue squamous carcinoma cells in vitro. This study laid a foundation for further investigation on the influence of the EMP1 gene on the metastasis of oral tongue squamous carcinoma and its molecular mechanism.
.</p>


Subject(s)
Carcinoma, Squamous Cell , Cell Line, Tumor , Cell Movement , Eukaryotic Cells , Genetic Vectors , Green Fluorescent Proteins , Humans , Neoplasm Proteins , Plasmids , Receptors, Cell Surface , Tongue Neoplasms , Transfection
10.
Article in English | WPRIM | ID: wpr-121455

ABSTRACT

Cellular prion protein (PrP(C)) is ubiquitously expressed in the cytomembrane of a considerable number of eukaryotic cells. Although several studies have investigated the functions of PrP(C) in cell proliferation, cell apoptosis, and tumorigenesis of mammals, the correlated functions of chicken PrP(C) (chPrP(C)) remain unknown. In this study, stable chPrP(C)-downregulated Marek's disease (MD) virus-transformed avian T cells (MSB1-SiRNA-3) were established by introducing short interfering RNA (SiRNA) targeting chicken prion protein genes. We found that downregulation of chPrP(C) inhibits proliferation, invasion, and migration, and induces G1 cell cycle phase arrest and apoptosis of MSB1-SiRNA-3 cells compared with Marek's disease virus-transformed avian T cells (MSB1) and negative control cells. To the best of our knowledge, the present study provides the first evidence supporting the positive correlation between the expression level of chPrP(C) and the proliferation, migration, and invasion ability of MSB1 cells, but appears to protect MSB1 cells from apoptosis, which suggests it functions in the formation and development of MD tumors. This evidence may contribute to future research into the specific molecular mechanisms of chPrP(C) in the formation and development of MD tumors.


Subject(s)
Animals , Apoptosis , Carcinogenesis , Cell Cycle , Cell Proliferation , Chickens , Down-Regulation , Eukaryotic Cells , Mammals , Marek Disease , RNA, Small Interfering , T-Lymphocytes
11.
Article in English | WPRIM | ID: wpr-98412

ABSTRACT

Autophagy is a principle catabolic process mediated by lysosomes in eukaryotic cells. This is an intracellular homeostatic mechanism crucial for degradation in acidic lysosomal compartments of waste components from the cytoplasm. Autophagy research was initially focused on its degradation mechanism, but focus is now shifting to its effects on immunity. It contributes to detection and removal of pathogens as well as regulation of inflammasomes and neutrophil extracellular traps. Moreover, it is pivotal in antigen presentation and immune cell maturation, survival and homeostasis. The importance of autophagic pathways in normal and dysregulated immunity has become increasingly recognized in the past several years. Dysregulation of the autophagic pathway is implicated in the pathogenesis of several rheumatic diseases. In this review, we summarize the immunological function of autophagy in innate and adaptive immunity, and the functions of autophagy in the pathogenesis of rheumatic diseases.


Subject(s)
Adaptive Immunity , Antigen Presentation , Arthritis, Rheumatoid , Autophagy , Cytoplasm , Eukaryotic Cells , Extracellular Traps , Homeostasis , Inflammasomes , Lupus Erythematosus, Systemic , Lysosomes , Rheumatic Diseases
12.
Journal of Breast Cancer ; : 133-141, 2016.
Article in English | WPRIM | ID: wpr-166641

ABSTRACT

PURPOSE: The promoter methylation status of cell cycle regulatory genes plays a crucial role in the regulation of the eukaryotic cell cycle. CpG cytosines are actively subjected to methylation during tumorigenesis, resulting in gain/loss of function. E2F5 gene has growth repressive activities; various studies suggest its involvement in tumorigenesis. This study aims to investigate the epigenetic regulation of E2F5 in breast cancer to better understand tumor biology. METHODS: The promoter methylation status of 50 breast tumor tissues and adjacent normal control tissues was analyzed. mRNA expression was determined using SYBR® green quantitative polymerase chain reaction (PCR), and methylation-specific PCR was performed for bisulfite-modified genomic DNA using E2F5-specific primers to assess promoter methylation. Data was statistically analyzed. RESULTS: Significant (p<0.001) upregulation was observed in E2F5 expression among tumor tissues, relative to the control group. These samples were hypo-methylated at the E2F5 promoter region in the tumor tissues, compared to the control. Change in the methylation status (Δmeth) was significantly lower (p=0.022) in the tumor samples, indicating possible involvement in tumorigenesis. Patients at the postmenopausal stage showed higher methylation (75%) than those at the premenopausal stage (23.1%). Interestingly, methylation levels gradually increased from the early to the advanced stages of the disease (p<0.001), which suggests a putative role of E2F5 methylation in disease progression that can significantly modulate tumor biology at more advanced stage and at postmenopausal age (Pearson's r=0.99 and 0.86, respectively). Among tissues with different histological status, methylation frequency was higher in invasive lobular carcinoma (80.0%), followed by invasive ductal carcinoma (46.7%) and ductal carcinoma in situ (20.0%). CONCLUSION: Methylation is an important epigenetic factor that might be involved in the upregulation of E2F5 gene in tumor tissues, which can be used as a prognostic marker for breast cancer.


Subject(s)
Biology , Breast Neoplasms , Breast , Carcinogenesis , Carcinoma, Ductal , Carcinoma, Intraductal, Noninfiltrating , Carcinoma, Lobular , Cell Cycle , Disease Progression , DNA , E2F5 Transcription Factor , Epigenomics , Eukaryotic Cells , Genes, Regulator , Humans , Methylation , Polymerase Chain Reaction , Promoter Regions, Genetic , RNA, Messenger , Up-Regulation
13.
Article in English | WPRIM | ID: wpr-125855

ABSTRACT

Heat shock protein 90 (Hsp90) is an adenosine triphosphate dependent molecular chaperone in eukaryotic cells that regulates the activation and maintenance of numerous regulatory and signaling proteins including epidermal growth factor receptor, human epidermal growth factor receptor 2, mesenchymal-epithelial transition factor, cyclin-dependent kinase-4, protein kinase B, hypoxia-inducible factor 1alpha, and matrix metalloproteinase-2. Since many of Hsp90 clients are oncogenic proteins, Hsp90 has become an attractive therapeutic target for treatment of cancer. To discover small molecule inhibitors targeting Hsp90 chaperone machinery, several strategies have been employed, which results in three classes of inhibitors such as N-terminal inhibitors, C-terminal inhibitors, and inhibitors disrupting protein-protein interactions of Hsp90 chaperone machinery. Developing small molecule inhibitors that modulate protein-protein interactions of Hsp90 is a challenging task, although it offers many alternative opportunities for therapeutic intervention. The lack of well-defined binding pocket and starting points for drug design challenges medicinal chemists to discover small molecule inhibitors disrupting protein-protein interactions of Hsp90. The present review will focus on the current studies on small molecule inhibitors disrupting protein-protein interactions of Hsp90 chaperone machinery, provide biological background on the structure, function and mechanism of Hsp90's protein-protein interactions, and discuss the challenges and promise of its small molecule modulations.


Subject(s)
Adenosine Triphosphate , Drug Design , Eukaryotic Cells , Heat-Shock Proteins , Hot Temperature , Humans , Matrix Metalloproteinase 2 , Molecular Chaperones , Proto-Oncogene Proteins c-akt , ErbB Receptors
14.
Protein & Cell ; (12): 241-253, 2015.
Article in English | WPRIM | ID: wpr-757601

ABSTRACT

Cells can adapt to environment and development by reconstructing their transcriptional networks to regulate diverse cellular processes without altering the underlying DNA sequences. These alterations, namely epigenetic changes, occur during cell division, differentiation and cell death. Numerous evidences demonstrate that epigenetic changes are governed by various types of determinants, including DNA methylation patterns, histone posttranslational modification signatures, histone variants, chromatin remodeling, and recently discovered chromosome conformation characteristics and non-coding RNAs (ncRNAs). Here, we highlight recent efforts on how the two latter epigenetic factors participate in the sophisticated transcriptional process and describe emerging techniques which permit us to uncover and gain insights into the fascinating genomic regulation.


Subject(s)
Cell Death , Cell Differentiation , Cell Division , Chromatin , Chemistry , Metabolism , Chromatin Assembly and Disassembly , DNA Methylation , Epigenesis, Genetic , Eukaryotic Cells , Cell Biology , Metabolism , Histones , Genetics , Metabolism , Humans , Protein Processing, Post-Translational , RNA, Untranslated , Genetics , Metabolism , Transcription, Genetic
15.
Natural Product Sciences ; : 282-288, 2015.
Article in English | WPRIM | ID: wpr-7745

ABSTRACT

Viriditoxin is a fungal metabolite isolated from Paecilomyces variotii, which was derived from the giant jellyfish Nemopilema nomurai. Viriditoxin was reported to inhibit polymerization of FtsZ, which is a key protein for bacterial cell division and a structural homologue of eukaryotic tubulin. Both tubulin and FtsZ contain a GTP-binding domain, have GTPase activity, assemble into protofilaments, two-dimensional sheets, and protofilament rings, and share substantial structural identities. Accordingly, we hypothesized that viriditoxin may inhibit eukaryotic cell division by inhibiting tubulin polymerization as in the case of bacterial FtsZ inhibition. Docking simulation of viriditoxin to beta-tubulin indicated that it binds to the paclitaxel-binding domain and makes hydrogen bonds with Thr276 and Gly370 in the same manner as paclitaxel. Viriditoxin suppressed growth of A549 human lung cancer cells, and inhibited cell division with G2/M cell cycle arrest, leading to apoptotic cell death.


Subject(s)
Apoptosis , Cell Cycle Checkpoints , Cell Cycle , Cell Death , Cell Division , Eukaryotic Cells , GTP Phosphohydrolases , Humans , Hydrogen , Lung Neoplasms , Lung , Paclitaxel , Paecilomyces , Polymerization , Polymers , Tubulin
16.
Chinese Journal of Biotechnology ; (12): 1256-1265, 2014.
Article in Chinese | WPRIM | ID: wpr-345599

ABSTRACT

To establish a gene regulation system compatible with biopharmaceutical industry and gene therapy, we constructed a fusion protein of biotin ligase from Bacillus subtilis (BS-BirA) and the trans-activation domain, and used its expression vector as the regulatory vector. Meanwhile, BS-BirA-specific operators were ligated upstream of attenuated CMV promoter to obtain the response vector. In this way, a novel eukaryotic gene regulation system responsive to biotin was established and named BS-Biotin-On system. BS-Biotin-On system was further investigated with the enhancing green fluorescent protein (EGFP) as the reporter gene. The results showed that our system was superior to the current similar regulation system in its higher induction ratio, and that the expression of interest gene could be tuned in a rapid and efficient manner by changing the biotin concentrations in the cultures, Our results show that the established system may provide a new alternative for the exogenous gene modulation.


Subject(s)
Bacillus subtilis , Biotin , Chemistry , Eukaryotic Cells , Metabolism , Gene Expression Regulation , Genetic Vectors , Promoter Regions, Genetic , Trans-Activators
17.
Article in English | WPRIM | ID: wpr-60467

ABSTRACT

The kinesin superfamily is a class of motor proteins moving along microtubule filaments and playing essential roles in mitosis of eukaryotic cells. In the cancer biology, mitotic activity is an essential factor for development and metastasis of various cancers. Therefore, the inhibition of kinesin activity is suggested as an alternative cancer therapy. Accumulated clinical evidences have proved the potency of kinesin inhibitors in cancer treatments. In this review, we provided an overview of kinesins that play a critical role in the pathophysiology of various cancers and described the beneficial vs. side effects of their inhibitors that have been tested in both basic science and clinical studies.


Subject(s)
Biology , Eukaryotic Cells , Kinesin , Microtubules , Mitosis , Neoplasm Metastasis , Translational Medical Research
18.
Article in English | WPRIM | ID: wpr-16139

ABSTRACT

Non-thermal atmospheric-pressure plasma, also named cold plasma, is defined as a partly ionized gas. Therefore, it cannot be equated with plasma from blood; it is not biological in nature. Non-thermal atmospheric-pressure plasma is a new innovative approach in medicine not only for the treatment of wounds, but with a wide-range of other applications, as e.g. topical treatment of other skin diseases with microbial involvement or treatment of cancer diseases. This review emphasizes plasma effects on wound healing. Non-thermal atmospheric-pressure plasma can support wound healing by its antiseptic effects, by stimulation of proliferation and migration of wound relating skin cells, by activation or inhibition of integrin receptors on the cell surface or by its pro-angiogenic effect. We summarize the effects of plasma on eukaryotic cells, especially on keratinocytes in terms of viability, proliferation, DNA, adhesion molecules and angiogenesis together with the role of reactive oxygen species and other components of plasma. The outcome of first clinical trials regarding wound healing is pointed out.


Subject(s)
Cell Survival , DNA , Eukaryotic Cells , Keratinocytes , Plasma Gases , Plasma , Reactive Oxygen Species , Skin , Skin Diseases , Wound Healing , Wounds and Injuries
19.
Protein & Cell ; (12): 658-672, 2014.
Article in English | WPRIM | ID: wpr-757641

ABSTRACT

Eukaryotic membrane proteins, many of which are key players in various biological processes, constitute more than half of the drug targets and represent important candidates for structural studies. In contrast to their physiological significance, only very limited number of eukaryotic membrane protein structures have been obtained due to the technical challenges in the generation of recombinant proteins. In this review, we examine the major recombinant expression systems for eukaryotic membrane proteins and compare their relative advantages and disadvantages. We also attempted to summarize the recent technical strategies in the advancement of eukaryotic membrane protein purification and crystallization.


Subject(s)
Animals , Escherichia coli , Genetics , Eukaryotic Cells , Metabolism , Genetic Vectors , HEK293 Cells , Humans , Insecta , Cell Biology , Genetics , Membrane Proteins , Chemistry , Genetics , Metabolism , Recombinant Proteins , Chemistry , Metabolism , Yeasts , Genetics
20.
Razi Journal of Medical Sciences. 2013; 19 (104): 1-9
in Persian | IMEMR | ID: emr-127181

ABSTRACT

Cultivated mammalian cells, because of their capacity for proper protein folding, assembly and post-translational modification, have become the dominant system for production of recombinant proteins in clinical application. Therefore, the quality and efficacy of protein can be superior when expressed in mammalian cells compared to other hosts such as bacteria. Gene reporter systems have contributed greatly to the study of eukaryotic gene expression and regulation. Although reporter genes have played a significant role in numerous applications, both in vitro and in vivo, they are most frequently used as indicators of transcriptional activity in cells. Luciferase-reporter assays are widely used to monitor the cellular events related to transduction and gene expression regulated by specific cascades, such as PRL/Jak2/Stat 5 pathway. In this study, recombinant human growth hormone [rhGH] was produced in eukaryotic Chinese hamster ovary [CHO] cell and production and concentration of rhGH verified by ELISA and western blotting. Then, the biological activity of rhGH was assessed by a gene reporter assay system [containing LHRE, TK promoter and Luc gene], using HEK 293 cells transfected with GH receptor and response element for STAT-5 measuring luciferase activity on a Berthold luminometer. The date showed that rhGH could be produced by eukaryotic host in good quantities as assessed by ELISA and western blotting. The results of gene reporter assay showed that rhGH produced by CHO cells is able to induce GH intracellular cell signaling. The rhGH produced by CHO cells showed higher bioactivity when compared to commercial GH. rhGH could be produced in mammalian cell lines at high levels with higher bioactivity. Gene reporter assay is a sensitive, quantitative, rapid, easy, reproducible and safe system for assessment of bioactivity of recombinant proteins such as rhGH


Subject(s)
Animals , Cricetulus , Ovary , Eukaryotic Cells , Genes, Reporter , Enzyme-Linked Immunosorbent Assay , Blotting, Western
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