134 results on '"Kuprash DV"'
Search Results
2. A polymorphic microsatellite marker in the human p55 TNF receptor, CD120a
- Author
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Eskdale, J, Turestkaya, RL, Armstrong, C, Kuprash, DV, Nedospasov, SA, and Gallagher, G
- Published
- 2000
- Full Text
- View/download PDF
3. [The molecular cloning and characteristics of the chromosomal copy of the human gene coding for the tumor-necrosis factor receptor]
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Turetskaia, RL, Kuprash, DV, Udalova, IA, Azizov, MM, and Nedospasov, SA
- Published
- 2016
4. Cut from the same cloth: RNAs transcribed from regulatory elements.
- Author
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Stasevich EM, Simonova AV, Bogomolova EA, Murashko MM, Uvarova AN, Zheremyan EA, Korneev KV, Schwartz AM, Kuprash DV, and Demin DE
- Subjects
- Humans, Gene Expression Regulation, Enhancer Elements, Genetic, Promoter Regions, Genetic, MicroRNAs genetics, MicroRNAs metabolism, Neoplasms genetics, Animals, Transcription, Genetic
- Abstract
A certain degree of chromatin openness is necessary for the activity of transcription-regulating regions within the genome, facilitating accessibility to RNA polymerases and subsequent synthesis of regulatory element RNAs (regRNAs) from these regions. The rapidly increasing number of studies underscores the significance of regRNAs across diverse cellular processes and diseases, challenging the paradigm that these transcripts are non-functional transcriptional noise. This review explores the multifaceted roles of regRNAs in human cells, encompassing rather well-studied entities such as promoter RNAs and enhancer RNAs (eRNAs), while also providing insights into overshadowed silencer RNAs and insulator RNAs. Furthermore, we assess notable examples of shorter regRNAs, like miRNAs, snRNAs, and snoRNAs, playing important roles. Expanding our discourse, we deliberate on the potential usage of regRNAs as biomarkers and novel targets for cancer and other human diseases., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Stasevich E. M. reports financial support was provided by Russian Science Foundation. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)
- Published
- 2024
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5. Methods for Functional Characterization of Genetic Polymorphisms of Non-Coding Regulatory Regions of the Human Genome.
- Author
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Uvarova AN, Tkachenko EA, Stasevich EM, Zheremyan EA, Korneev KV, and Kuprash DV
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- Humans, Genome-Wide Association Study, Regulatory Sequences, Nucleic Acid, Computational Biology methods, Transcription Factors genetics, Transcription Factors metabolism, Genome, Human, Polymorphism, Genetic
- Abstract
Currently, numerous associations between genetic polymorphisms and various diseases have been characterized through the Genome-Wide Association Studies. Majority of the clinically significant polymorphisms are localized in non-coding regions of the genome. While modern bioinformatic resources make it possible to predict molecular mechanisms that explain influence of the non-coding polymorphisms on gene expression, such hypotheses require experimental verification. This review discusses the methods for elucidating molecular mechanisms underlying dependence of the disease pathogenesis on specific genetic variants within the non-coding sequences. A particular focus is on the methods for identification of transcription factors with binding efficiency dependent on polymorphic variations. Despite remarkable progress in bioinformatic resources enabling prediction of the impact of polymorphisms on the disease pathogenesis, there is still the need for experimental approaches to investigate this issue.
- Published
- 2024
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6. Reverse Genetics Applied to Immunobiology of Tumor Necrosis Factor, a Multifunctional Cytokine.
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Nedospasov SA, Kruglov AA, Tumanov AV, Drutskaya MS, Astrakhantseva IV, and Kuprash DV
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- Animals, Humans, Mice, Gene Editing methods, Neoplasms immunology, Neoplasms genetics, Neoplasms therapy, Tumor Necrosis Factor-alpha genetics, Tumor Necrosis Factor-alpha metabolism
- Abstract
Tumor necrosis factor (TNF) is one of many cytokines - protein molecules responsible for communication between the cells of immune system. TNF was discovered and given its grand name because of its striking antitumor effects in experimental systems, but its main physiological functions in the context of whole organism turned out to be completely unrelated to protection against tumors. This short review discusses "man-made" mouse models generated by early genome-editing technologies, which enabled us to establish true functions of TNF in health and certain diseases as well as to unravel potential strategies for improving therapy of TNF-dependent diseases.
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- 2024
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7. Breg-Mediated Immunoregulation in the Skin.
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Zheremyan EA, Ustiugova AS, Karamushka NM, Uvarova AN, Stasevich EM, Bogolyubova AV, Kuprash DV, and Korneev KV
- Subjects
- Humans, Skin, Wound Healing, B-Lymphocytes, Regulatory, Psoriasis, Dermatitis, Atopic
- Abstract
Wound healing is a complex process involving a coordinated series of events aimed at restoring tissue integrity and function. Regulatory B cells (Bregs) are a subset of B lymphocytes that play an essential role in fine-tuning immune responses and maintaining immune homeostasis. Recent studies have suggested that Bregs are important players in cutaneous immunity. This review summarizes the current understanding of the role of Bregs in skin immunity in health and pathology, such as diabetes, psoriasis, systemic sclerosis, cutaneous lupus erythematosus, cutaneous hypersensitivity, pemphigus, and dermatomyositis. We discuss the mechanisms by which Bregs maintain tissue homeostasis in the wound microenvironment through the promotion of angiogenesis, suppression of effector cells, and induction of regulatory immune cells. We also mention the potential clinical applications of Bregs in promoting wound healing, such as the use of adoptive Breg transfer.
- Published
- 2024
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8. rs71327024 Associated with COVID-19 Hospitalization Reduces CXCR6 Promoter Activity in Human CD4 + T Cells via Disruption of c-Myb Binding.
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Uvarova AN, Stasevich EM, Ustiugova AS, Mitkin NA, Zheremyan EA, Sheetikov SA, Zornikova KV, Bogolyubova AV, Rubtsov MA, Kulakovskiy IV, Kuprash DV, Korneev KV, and Schwartz AM
- Subjects
- Humans, Hospitalization, Promoter Regions, Genetic, Receptors, CXCR6 genetics, SARS-CoV-2, T-Lymphocytes, Helper-Inducer, COVID-19 genetics
- Abstract
Single-nucleotide polymorphism rs71327024 located in the human 3p21.31 locus has been associated with an elevated risk of hospitalization upon SARS-CoV-2 infection. The 3p21.31 locus contains several genes encoding chemokine receptors potentially relevant to severe COVID-19. In particular, CXCR6, which is prominently expressed in T lymphocytes, NK, and NKT cells, has been shown to be involved in the recruitment of immune cells to non-lymphoid organs in chronic inflammatory and respiratory diseases. In COVID-19, CXCR6 expression is reduced in lung resident memory T cells from patients with severe disease as compared to the control cohort with moderate symptoms. We demonstrate here that rs71327024 is located within an active enhancer that augments the activity of the CXCR6 promoter in human CD4
+ T lymphocytes. The common rs71327024(G) variant makes a functional binding site for the c-Myb transcription factor, while the risk rs71327024(T) variant disrupts c-Myb binding and reduces the enhancer activity. Concordantly, c-Myb knockdown in PMA-treated Jurkat cells negates rs71327024's allele-specific effect on CXCR6 promoter activity. We conclude that a disrupted c-Myb binding site may decrease CXCR6 expression in T helper cells of individuals carrying the minor rs71327024(T) allele and thus may promote the progression of severe COVID-19 and other inflammatory pathologies.- Published
- 2023
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9. Differentially activated B cells develop regulatory phenotype and show varying immunosuppressive features: a comparative study.
- Author
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Zheremyan EA, Ustiugova AS, Uvarova AN, Karamushka NM, Stasevich EM, Gogoleva VS, Bogolyubova AV, Mitkin NA, Kuprash DV, and Korneev KV
- Subjects
- Humans, Immunosuppressive Agents pharmacology, Immunosuppressive Agents therapeutic use, Immunosuppression Therapy, Phenotype, CD40 Ligand, B-Lymphocytes, Regulatory
- Abstract
Regulatory B lymphocytes (Bregs) are B cells with well-pronounced immunosuppressive properties, allowing them to suppress the activity of effector cells. A broad repertoire of immunosuppressive mechanisms makes Bregs an attractive tool for adoptive cell therapy for diseases associated with excessive activation of immune reactions. Such therapy implies Breg extraction from the patient's peripheral blood, ex vivo activation and expansion, and further infusion into the patient. At the same time, the utility of Bregs for therapeutic approaches is limited by their small numbers and extremely low survival rate, which is typical for all primary B cell cultures. Therefore, extracting CD19
+ cells from the patient's peripheral blood and specifically activating them ex vivo to make B cells acquire a suppressive phenotype seems to be far more productive. It will allow a much larger number of B cells to be obtained initially, which may significantly increase the likelihood of successful immunosuppression after adoptive Breg transfer. This comparative study focuses on finding ways to efficiently manipulate B cells in vitro to differentiate them into Bregs. We used CD40L, CpG, IL4, IL21, PMA, and ionomycin in various combinations to generate immunosuppressive phenotype in B cells and performed functional assays to test their regulatory capacity. This work shows that treatment of primary B cells using CD40L + CpG + IL21 mix was most effective in terms of induction of functionally active regulatory B lymphocytes with high immunosuppressive capacity ex vivo ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Zheremyan, Ustiugova, Uvarova, Karamushka, Stasevich, Gogoleva, Bogolyubova, Mitkin, Kuprash and Korneev.)- Published
- 2023
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10. Adversary of DNA integrity: A long non-coding RNA stimulates driver oncogenic chromosomal rearrangement in human thyroid cells.
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Demin DE, Murashko MM, Uvarova AN, Stasevich EM, Shyrokova EY, Gorlachev GE, Zaretsky AR, Korneev KV, Ustiugova AS, Tkachenko EA, Kostenko VV, Tatosyan KA, Sheetikov SA, Spirin PV, Kuprash DV, and Schwartz AM
- Subjects
- Humans, Thyroid Cancer, Papillary genetics, Chromosome Aberrations, Gene Rearrangement, Carcinogenesis genetics, RNA, Long Noncoding genetics, Thyroid Neoplasms pathology
- Abstract
The flurry of publications devoted to the functions of long non-coding RNAs (lncRNAs) published in the last decade leaves no doubt about the exceptional importance of lncRNAs in various areas including tumor biology. However, contribution of lncRNAs to the early stages of oncogenesis remains poorly understood. In this study we explored a new role for lncRNAs: stimulation of specific chromosomal rearrangements upon DNA damage. We demonstrated that lncRNA CASTL1 (ENSG00000269945) stimulates the formation of the CCDC6-RET inversion (RET/PTC1) in human thyroid cells subjected to radiation or chemical DNA damage. Facilitation of chromosomal rearrangement requires lncRNA to contain regions complementary to the introns of both CCDC6 and RET genes as deletion of these regions deprives CASTL1 of the ability to stimulate the gene fusion. We found that CASTL1 expression is elevated in tumors with CCDC6-RET fusion which is the most frequent rearrangement in papillary thyroid carcinoma. Our results open a new venue for the studies of early oncogenesis in various tumor types, especially those associated with physical or chemical DNA damage., (© 2022 UICC.)
- Published
- 2023
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11. Corrigendum to "CRISPR/Cas9 genome editing demonstrates functionality of the autoimmunity-associated SNP rs12946510" [Biochim. Biophys. Acta (BBA) - Mol. Basis Dis. 1869 (2023) 166599].
- Author
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Ustiugova AS, Dvorianinova EM, Melnikova NV, Dmitriev AA, Kuprash DV, and Afanasyeva MA
- Abstract
Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
- Published
- 2023
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12. Interaction Between Adipocytes and B Lymphocytes in Human Metabolic Diseases.
- Author
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Stasevich EM, Zheremyan EA, Kuprash DV, and Schwartz AM
- Subjects
- Humans, Adipocytes metabolism, Adipose Tissue metabolism, Glucose metabolism, Inflammation metabolism, B-Lymphocytes metabolism, Diabetes Mellitus, Type 2 metabolism, Metabolic Diseases pathology
- Abstract
Diseases associated with the disorders of carbohydrate and lipid metabolism are widespread in the modern world. Interaction between the cells of adipose tissue - adipocytes - and immune system cells is an essential factor in pathogenesis of such diseases. Long-term increase in the glucose and fatty acid levels leads to adipocyte hypertrophy and increased expression of pro-inflammatory cytokines and adipokines by these cells. As a result, immune cells acquire a pro-inflammatory phenotype, and new leukocytes are recruited. Inflammation of adipose tissue leads to insulin resistance and stimulates formation of atherosclerotic plaques and development of autoimmunity. New studies show that different groups of B lymphocytes play an essential role in regulation of adipose tissue inflammation. Decrease in the number of B-2 lymphocytes suppresses development of a number of metabolic diseases, whereas decreased numbers of the regulatory B lymphocytes and B-1 lymphocytes are associated with more severe pathology. Recent studies showed that adipocytes influence B lymphocyte activity both directly and by altering activity of other immune cells. These findings provide better understanding of the molecular mechanisms of human pathologies associated with impaired carbohydrate and lipid metabolism, such as type 2 diabetes mellitus.
- Published
- 2023
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13. Novel Potential Mechanisms of Regulatory B Cell-Mediated Immunosuppression.
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Zheremyan EA, Ustiugova AS, Radko AI, Stasevich EM, Uvarova AN, Mitkin NA, Kuprash DV, and Korneev KV
- Subjects
- Humans, Immune Tolerance, Immunosuppressive Agents metabolism, Immunosuppression Therapy, L-Amino Acid Oxidase metabolism, B-Lymphocytes, Regulatory metabolism, B-Lymphocytes, Regulatory pathology
- Abstract
B lymphocytes play an important role in the regulation of immune response in both normal and pathological conditions. Traditionally, the main functions of B cells were considered to be antibody production and antigen presentation, but in recent decades there have been discovered several subpopulations of regulatory B lymphocytes (Bregs), which maintain immunological tolerance and prevent overactivation of the immune system. Memory (mBregs, CD19
+ CD24hi CD27+ ) and transitional (tBregs, CD19+ CD24hi CD38hi ) subpopulations of Bregs are usually considered in the context of studying the role of these B cells in various human pathologies. However, the mechanisms by which these Breg subpopulations exert their immunosuppressive activity remain poorly understood. In this work, we used bioinformatic analysis of open-source RNA sequencing data to propose potential mechanisms of B cell-mediated immunosuppression. Analysis of differential gene expression before and after activation of these subpopulations allowed us to identify six candidate molecules that may determine the functionality of mBregs and tBregs. IL4I1-, SIRPA-, and SLAMF7-dependent mechanisms of immunosuppression may be characteristic of both Breg subsets, while NID1-, CST7-, and ADORA2B-dependent mechanisms may be predominantly characteristic of tBregs. In-depth understanding of the molecular mechanisms of anti-inflammatory immune response of B lymphocytes is an important task for both basic science and applied medicine and could facilitate the development of new approaches to the therapy of complex diseases.- Published
- 2023
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14. LTα, TNF, and ILC3 in Peyer's Patch Organogenesis.
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Gogoleva VS, Kuprash DV, Grivennikov SI, Tumanov AV, Kruglov AA, and Nedospasov SA
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- Animals, Lymphoid Tissue, Mice, Mice, Knockout, Organogenesis genetics, Tumor Necrosis Factors metabolism, Lymphotoxin-alpha, Peyer's Patches
- Abstract
TNF and LTα are structurally related cytokines of the TNF superfamily. Their genes are located in close proximity to each other and to the Ltb gene within the TNF/LT locus inside MHC. Unlike Ltb , transcription of Tnf and of Lta is tightly controlled, with the Tnf gene being an immediate early gene that is rapidly induced in response to various inflammatory stimuli. Genes of the TNF/LT locus play a crucial role in lymphoid tissue organogenesis, although some aspects of their specific contribution remain controversial. Here, we present new findings and discuss the distinct contribution of TNF produced by ILC3 cells to Peyer's patch organogenesis.
- Published
- 2022
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15. Enhancer RNA AL928768.3 from the IGH Locus Regulates MYC Expression and Controls the Proliferation and Chemoresistance of Burkitt Lymphoma Cells with IGH/MYC Translocation.
- Author
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Stasevich EM, Uvarova AN, Murashko MM, Khabusheva ER, Sheetikov SA, Prassolov VS, Kuprash DV, Demin DE, and Schwartz AM
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- Cell Proliferation, Drug Resistance, Neoplasm genetics, Humans, RNA, Translocation, Genetic, Burkitt Lymphoma drug therapy, Burkitt Lymphoma genetics, Burkitt Lymphoma pathology, Lymphoma genetics
- Abstract
Chromosomal rearrangements leading to the relocation of proto-oncogenes into transcription-active regions are found in various types of tumors. In particular, the transfer of proto-oncogenes to the locus of heavy chains of immunoglobulins (IGH) is frequently observed in B-lymphomas. The increased expression of the MYC proto-oncogene due to IGH/MYC translocation is detected in approximately 85% of Burkitt lymphoma cases. The regulatory mechanisms affecting the oncogenes upon translocation include non-coding enhancer RNAs (eRNAs). We conducted a search for the eRNAs that may affect MYC transcription in the case of IGH/MYC translocation in Burkitt lymphoma, looking for potentially oncogenic eRNAs located at the IGH locus and predominantly expressed in B cells. Overexpression and knockdown of our primary candidate eRNA AL928768.3 led to the corresponding changes in the expression of MYC proto-oncogene in Burkitt lymphoma cells. Furthermore, we demonstrated that AL928768.3 knockdown decreased lymphoma cell proliferation and resistance to chemotherapy. Significant effects were observed only in cell lines bearing IGH/MYC abnormality but not in B-cell lines without this translocation nor primary B-cells. Our results indicate that AL928768.3 plays an important role in the development of Burkitt's lymphoma and suggest it and similar, yet undiscovered eRNAs as potential tissue-specific targets for cancer treatment.
- Published
- 2022
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16. [The Minor T Allele of the Single Nucleotide Polymorphism rs 13360222 Decreases the Activity of the HAVCR2 Gene Enhancer in a Cell Model of Human Macrophages].
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Uvarova AN, Ustiugova AS, Mitkin NA, Schwartz AM, Korneev KV, and Kuprash DV
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- Alleles, Humans, Introns, Promoter Regions, Genetic, U937 Cells, Hepatitis A Virus Cellular Receptor 2 genetics, Macrophages, Polymorphism, Single Nucleotide
- Abstract
The TIM-3 receptor, encoded by the Hepatitis A Virus Cellular Receptor 2 (HAVCR2) gene, is an immune checkpoint and plays an important role in preventing the development of autoimmune reactions. This receptor is expressed on the surface of various immunocytes and its functions in myeloid cells remain poorly understood, compared to the role of T cell specific TIM-3 that is actively studied in the context of the search for promising therapeutic targets in cancer immunotherapy. During this study, we performed deletion analysis of the promoter region of the HAVCR2 gene, as well as functional characterization of its enhancer, and studied the effect of a number of single nucleotide polymorphisms (SNPs) on the activity of these regulatory elements in the relevant model of human macrophage-like cells-U937 activated monocytes. We have shown that the SNPs rs10515746(A) and rs4704853(A) located in the HAVCR2 gene promoter and associated with the development of a number of pathologies, do not affect the activity of the promoter in activated monocytes. However, a minor T variant of SNP rs13360222 located in the enhancer in the third intron of the gene, significantly reduces the ability of the enhancer to activate the HAVCR2 promoter, presumably due to weakening of the binding of nuclear receptor ESR2 to the respective region.
- Published
- 2022
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17. EGR1 and RXRA transcription factors link TGF-β pathway and CCL2 expression in triple negative breast cancer cells.
- Author
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Gorbacheva AM, Uvarova AN, Ustiugova AS, Bhattacharyya A, Korneev KV, Kuprash DV, and Mitkin NA
- Subjects
- Base Sequence, Binding Sites, Cell Line, Tumor, Chemokine CCL2 genetics, Female, Gene Expression Regulation, Neoplastic, Humans, Models, Biological, Point Mutation genetics, Promoter Regions, Genetic genetics, Protein Binding, Triple Negative Breast Neoplasms genetics, Triple Negative Breast Neoplasms pathology, Chemokine CCL2 metabolism, Early Growth Response Protein 1 metabolism, Retinoid X Receptor alpha metabolism, Signal Transduction, Transforming Growth Factor beta metabolism, Triple Negative Breast Neoplasms metabolism
- Abstract
Transforming growth factor beta (TGF-β) is the main cytokine responsible for the induction of the epithelial-mesenchymal transition of breast cancer cells, which is a hallmark of tumor transformation to the metastatic phenotype. Recently, research demonstrated that the chemokine CCL2 gene expression level directly correlates with the TGF-β activity in breast cancer patients. CCL2 attracts tumor-associated macrophages and is, therefore, considered as an important inductor of breast cancer progression; however, the precise mechanisms underlying its regulation by TGF-β are unknown. Here, we studied the behavior of the CCL2 gene in MDA-MB-231 and HCC1937 breast cancer cells representing mesenchymal-like phenotype activated by TGF-β. Using bioinformatics, deletion screening and point mutagenesis, we identified binding sites in the CCL2 promoter and candidate transcription factors responsible for its regulation by TGF-β. Among these factors, only the knock-down of EGR1 and RXRA made CCL2 promoter activity independent of TGF-β. These factors also demonstrated binding to the CCL2 promoter in a TGF-β-dependent manner in a chromatin immunoprecipitation assay, and point mutations in the EGR1 and RXRA binding sites totally abolished the effect of TGF-β. Our results highlight the key role of EGR1 and RXRA transcription factors in the regulation of CCL2 gene in response to TGF-β pathway.
- Published
- 2021
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18. The Role of RNA in DNA Breaks, Repair and Chromosomal Rearrangements.
- Author
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Murashko MM, Stasevich EM, Schwartz AM, Kuprash DV, Uvarova AN, and Demin DE
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- Animals, DNA Breaks, Double-Stranded, Humans, MicroRNAs genetics, Chromosome Aberrations, DNA Repair, MicroRNAs metabolism
- Abstract
Incorrect reparation of DNA double-strand breaks (DSB) leading to chromosomal rearrangements is one of oncogenesis's primary causes. Recently published data elucidate the key role of various types of RNA in DSB formation, recognition and repair. With growing interest in RNA biology, increasing RNAs are classified as crucial at the different stages of the main pathways of DSB repair in eukaryotic cells: nonhomologous end joining (NHEJ) and homology-directed repair (HDR). Gene mutations or variation in expression levels of such RNAs can lead to local DNA repair defects, increasing the chromosome aberration frequency. Moreover, it was demonstrated that some RNAs could stimulate long-range chromosomal rearrangements. In this review, we discuss recent evidence demonstrating the role of various RNAs in DSB formation and repair. We also consider how RNA may mediate certain chromosomal rearrangements in a sequence-specific manner.
- Published
- 2021
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19. [Regulation of IL33 Gene Expression by SP1 and Foxa1 in Breast and Lung Cancer Cells].
- Author
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Gorbacheva AM, Kuprash DV, and Mitkin NA
- Subjects
- Cell Line, Tumor, Gene Expression, Gene Expression Regulation, Neoplastic, Hepatocyte Nuclear Factor 3-alpha genetics, Humans, Interleukin-33 genetics, Sp1 Transcription Factor genetics, Tumor Microenvironment, Breast Neoplasms genetics, Lung Neoplasms genetics
- Abstract
Interleukin-33 (IL-33) is a member of the IL-1 cytokine family, primarily known as a mediator of the humoral immune response. It provides protection of barrier tissues and participates in the development of a range of diseases. This cytokine promotes carcinogenesis by induction of proliferation and survival of cancer cells, remodeling of the tumor microenvironment, and promoting immunosuppressive conditions. Elevated levels of IL-33 were observed in many types of cancers. This elevation correlates with a poor prognosis, making IL33 a promising target for cancer immunotherapy. The mechanisms of IL-33 expression regulation in human tumor cells are not well understood. Here, we show that that expression of IL-33 in breast and lung cancer cell lines depends, at least in part, on the activity of the SP1 and FOXA1 transcription factors. Increases in the activity of these transcription factors may be responsible for elevated levels of IL-33 and subsequent tumor progression.
- Published
- 2021
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20. Minor C allele of the SNP rs7873784 associated with rheumatoid arthritis and type-2 diabetes mellitus binds PU.1 and enhances TLR4 expression.
- Author
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Korneev KV, Sviriaeva EN, Mitkin NA, Gorbacheva AM, Uvarova AN, Ustiugova AS, Polanovsky OL, Kulakovskiy IV, Afanasyeva MA, Schwartz AM, and Kuprash DV
- Subjects
- 3' Untranslated Regions genetics, Alleles, Cell Line, Cell Line, Tumor, HEK293 Cells, Humans, Promoter Regions, Genetic genetics, U937 Cells, Arthritis, Rheumatoid genetics, Diabetes Mellitus, Type 2 genetics, Genetic Predisposition to Disease genetics, Polymorphism, Single Nucleotide genetics, Proto-Oncogene Proteins genetics, Toll-Like Receptor 4 genetics, Trans-Activators genetics
- Abstract
Toll-like receptor 4 (TLR4) is an innate immunity receptor predominantly expressed on myeloid cells and involved in the development of various diseases, many of them with complex genetics. Here we present data on functionality of single nucleotide polymorphism rs7873784 located in the 3'-untranslated region (3'-UTR) of TLR4 gene and associated with various pathologies involving chronic inflammation. We demonstrate that TLR4 3'-UTR strongly enhanced the activity of TLR4 promoter in U937 human monocytic cell line while minor rs7873784(C) allele created a binding site for transcription factor PU.1 (encoded by SPI1 gene), a known regulator of TLR4 expression. Increased binding of PU.1 further augmented the TLR4 transcription while PU.1 knockdown or complete disruption of the PU.1 binding site abrogated the effect. We hypothesize that additional functional PU.1 site may increase TLR4 expression in individuals carrying minor C variant of rs7873784 and modulate the development of certain pathologies, such as rheumatoid arthritis and type-2 diabetes mellitus., (Copyright © 2019 Elsevier B.V. All rights reserved.)
- Published
- 2020
- Full Text
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21. Multi-dimensional immunoproteomics coupled with in vitro recapitulation of oncogenic NRAS Q61R identifies diagnostically relevant autoantibody biomarkers in thyroid neoplasia.
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Belousov PV, Afanasyeva MA, Gubernatorova EO, Bogolyubova AV, Uvarova AN, Putlyaeva LV, Ramanauskaite EM, Kopylov AT, Demin DE, Tatosyan KA, Ustiugova AS, Prokofjeva MM, Lanshchakov KV, Vanushko VE, Zaretsky AR, Severskaia NV, Dvinskikh NY, Abrosimov AY, Kuprash DV, and Schwartz AM
- Subjects
- Biomarkers, Tumor metabolism, Case-Control Studies, Cell Line, Tumor, Early Detection of Cancer, Female, GTP Phosphohydrolases immunology, Humans, Membrane Proteins immunology, Mutation, Thyroid Neoplasms immunology, Autoantibodies metabolism, GTP Phosphohydrolases genetics, Membrane Proteins genetics, Proteomics methods, Thyroid Neoplasms diagnosis
- Abstract
Tumor-associated antigen (TAA)-specific autoantibodies have been widely implicated in cancer diagnosis. However, cancer cell lines that are typically exploited as candidate TAA sources in immunoproteomic studies may fail to accurately represent the autoantigen-ome of lower-grade neoplasms. Here, we established an integrated strategy for the identification of disease-relevant TAAs in thyroid neoplasia, which combined NRAS
Q61R oncogene expression in non-tumorous thyroid Nthy-ori 3-1 cells with a multi-dimensional proteomic technique DISER that consisted of profiling NRASQ61R -induced proteins using 2-dimensional difference gel electrophoresis (2D-DIGE) coupled with serological proteome analysis (SERPA) of the TAA repertoire of patients with thyroid encapsulated follicular-patterned/RAS-like phenotype (EFP/RLP) tumors. We identified several candidate cell-based (nicotinamide phosphoribosyltransferase NAMPT, glutamate dehydrogenase GLUD1, and glutathione S-transferase omega-1 GSTO1) and autoantibody (fumarate hydratase FH, calponin-3 CNN3, and pyruvate kinase PKM autoantibodies) biomarkers, including NRASQ61R -induced TAA phosphoglycerate kinase 1 PGK1. Meta-profiling of the reactivity of the identified autoantibodies across an independent SERPA series implicated the PKM autoantibody as a histological phenotype-independent biomarker of thyroid malignancy (11/38 (29%) patients with overtly malignant and uncertain malignant potential (UMP) tumors vs 0/22 (p = 0.0046) and 0/20 (p = 0.011) patients with non-invasive EFP/RLP tumors and healthy controls, respectively). PGK1 and CNN3 autoantibodies were identified as EFP/RLP-specific biomarkers, potentially suitable for further discriminating tumors with different malignant potential (PGK1: 7/22 (32%) patients with non-invasive EFP/RLP tumors vs 0/38 (p = 0.00044) and 0/20 (p = 0.0092) patients with other tumors and healthy controls, respectively; СNN3: 9/29 (31%) patients with malignant and borderline EFP/RLP tumors vs 0/31 (p = 0.00068) and 0/20 (p = 0.0067) patients with other tumors and healthy controls, respectively). The combined use of PKM, CNN3, and PGK1 autoantibodies allowed the reclassification of malignant/UMP tumor risk in 19/41 (46%) of EFP/RLP tumor patients. Taken together, we established an experimental pipeline DISER for the concurrent identification of cell-based and TAA biomarkers. The combination of DISER with in vitro oncogene expression allows further targeted identification of oncogene-induced TAAs. Using this integrated approach, we identified candidate autoantibody biomarkers that might be of value for differential diagnostic purposes in thyroid neoplasia., (Copyright © 2019 Elsevier B.V. All rights reserved.)- Published
- 2019
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22. [Relative Efficiency of Transcription Factor Binding to Allelic Variants of Regulatory Regions of Human Genes in Immunoprecipitation and Real-Time PCR].
- Author
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Mitkin NA, Korneev KV, Gorbacheva AM, and Kuprash DV
- Subjects
- Humans, Protein Binding, Alleles, Immunoprecipitation, Polymorphism, Single Nucleotide, Real-Time Polymerase Chain Reaction, Regulatory Sequences, Nucleic Acid genetics, Transcription Factors metabolism
- Abstract
The efficiency at which specific transcription factors interact with DNA may vary in the presence of single nucleotide polymorphisms (SNPs), and the variation provides an important mechanism that regulates expression of human genes and contributes to the individual susceptibility to various diseases. Ample genetic and epigenetic data make it possible to predict both functional polymorphic variants and the transcription factors whose binding they affect. However, predictions of the kind require experimental verification. An original method developed for the purpose includes immunoprecipitation of DNA-protein complexes, followed by quantification of the bound DNA by real-time PCR. The method does not require chemical modification of the DNA probes and yield reproducible results with total nuclear extracts from cultured human cells.
- Published
- 2019
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23. Functional SNPs in the Human Autoimmunity-Associated Locus 17q12-21.
- Author
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Ustiugova AS, Korneev KV, Kuprash DV, and Afanasyeva AMA
- Subjects
- Forkhead Box Protein O1 genetics, Forkhead Box Protein O1 metabolism, Humans, Jurkat Cells, Linkage Disequilibrium, MEF2 Transcription Factors genetics, MEF2 Transcription Factors metabolism, Quantitative Trait Loci, Autoimmune Diseases genetics, Chromosomes, Human, Pair 17 genetics, Polymorphism, Single Nucleotide
- Abstract
Genome-wide association studies (GWASes) revealed several single-nucleotide polymorphisms (SNPs) in the human 17q12-21 locus associated with autoimmune diseases. However, follow-up studies are still needed to identify causative SNPs directly mediating autoimmune risk in the locus. We have chosen six SNPs in high linkage disequilibrium with the GWAS hits that showed the strongest evidence of causality according to association pattern and epigenetic data and assessed their functionality in a local genomic context using luciferase reporter system. We found that rs12946510, rs4795397, rs12709365, and rs8067378 influenced the reporter expression level in leukocytic cell lines. The strongest effect visible in three distinct cell types was observed for rs12946510 that is predicted to alter MEF2A/C and FOXO1 binding sites., Competing Interests: The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.
- Published
- 2019
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24. Glucocorticoid Receptor Binding Inhibits an Intronic IL33 Enhancer and is Disrupted by rs4742170 (T) Allele Associated with Specific Wheezing Phenotype in Early Childhood.
- Author
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Gorbacheva AM, Kuprash DV, and Mitkin NA
- Subjects
- Base Sequence, Binding Sites, Cell Line, Tumor, Child, Preschool, Humans, Hydrocortisone pharmacology, Phenotype, Phosphorylation drug effects, Promoter Regions, Genetic, Alleles, Enhancer Elements, Genetic genetics, Interleukin-33 genetics, Introns genetics, Polymorphism, Single Nucleotide genetics, Receptors, Glucocorticoid metabolism, Respiratory Sounds genetics
- Abstract
Interleukin 33 (IL-33) is a cytokine constitutively expressed by various cells of barrier tissues that contribute to the development of inflammatory immune responses. According to its function as an alarmin secreted by lung and airway epithelium, IL-33 plays a significant role in pathogenesis of allergic disorders. IL-33 is strongly involved in the pathogenesis of asthma, anaphylaxis, allergy and dermatitis, and genetic variations in IL33 locus are associated with increased susceptibility to asthma. Genome-wide association studies have identified risk "T" allele of the single-nucleotide polymorphism rs4742170 located in putative IL33 enhancer area as susceptible variant for development of specific wheezing phenotype in early childhood. Here, we demonstrate that risk "T" rs4742170 allele disrupts binding of glucocorticoid receptor (GR) transcription factor to IL33 putative enhancer. The IL33 promoter/enhancer constructs containing either 4742170 (T) allele or point mutations in the GR-binding site, were significantly more active and did not respond to cortisol in a pulmonary epithelial cell line. At the same time, the constructs containing rs4742170 (C) allele with a functional GR-binding site were less active and further inhibitable by cortisol. The latter effect was GR-dependent as it was completely abolished by GR-specific siRNA. This mechanism may explain the negative effect of the rs4742170 (T) risk allele on the development of wheezing phenotype that strongly correlates with allergic sensitization in childhood.
- Published
- 2018
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25. Potential Markers of Autoimmune Diseases, Alleles rs115662534(T) and rs548231435(C), Disrupt the Binding of Transcription Factors STAT1 and EBF1 to the Regulatory Elements of Human CD40 Gene.
- Author
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Putlyaeva LV, Demin DE, Korneev KV, Kasyanov AS, Tatosyan KA, Kulakovskiy IV, Kuprash DV, and Schwartz AM
- Subjects
- Base Sequence, Biomarkers metabolism, Cell Line, Tumor, Gene Expression Regulation, Neoplastic genetics, Genes, Reporter genetics, Humans, Introns genetics, Protein Binding, Alleles, Autoimmune Diseases genetics, CD40 Antigens genetics, Enhancer Elements, Genetic genetics, Polymorphism, Single Nucleotide, STAT1 Transcription Factor metabolism, Trans-Activators metabolism
- Abstract
CD40 receptor is expressed on B lymphocytes and other professional antigen-presenting cells. The binding of CD40 to its ligand CD154 on the surface of T helper cells plays an important role in the activation of B lymphocytes required for production of antibodies, in particular, against autoantigens. Association of several single nucleotide polymorphisms (SNPs) located in the non-coding areas of human CD40 locus with the elevated risk of autoimmune diseases has been demonstrated. The most studied of these SNPs is rs4810485 located in the first intron of the CD40 gene. Expression of the CD40 gene in B lymphocytes of donors homozygous for the common allelic variant of this polymorphism (G) is higher than in B cells from donors carrying the minor (T) variant. We investigated the enhancer activity of this fragment of the CD40 locus in human B cell lines and showed that it is independent on the rs4810485 alleles. However, the minor allelic variants of the rs4810485-linked SNPs rs548231435 and rs115662534 were associated with a significant decrease in the activity of the CD40 promoter due to the impairments in the binding of EBF1 and STAT1 transcription factors, respectively.
- Published
- 2018
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26. Neonatal Lethality and Inflammatory Phenotype of the New Transgenic Mice with Overexpression of Human Interleukin-6 in Myeloid Cells.
- Author
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Zvartsev RV, Korshunova DS, Gorshkova EA, Nosenko MA, Korneev KV, Maksimenko OG, Korobko IV, Kuprash DV, Drutskaya MS, Nedospasov SA, and Deikin AV
- Subjects
- Animals, Green Fluorescent Proteins biosynthesis, Green Fluorescent Proteins genetics, Humans, Macrophages cytology, Mice, Mice, Transgenic, Monocytes cytology, Hematopoiesis, Interleukin-6 biosynthesis, Interleukin-6 genetics, Macrophages metabolism, Monocytes metabolism
- Abstract
To model human interleukin-6 (hIL-6) associated diseases, unique mice with transgenic overexpression of human IL-6 and reporter fluorescent protein EGFP in cells of macrophage-monocyte lineage were generated using loxP-Cre system. High level of hIL-6 production by macrophages and monocytes, as confirmed in vitro in primary culture of bone marrow-derived macrophages, in vivo resulted in early postnatal death in vivo, presumably, due to the effect of overexpression of hIL-6 on hematopoiesis.
- Published
- 2018
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27. Protective C allele of the single-nucleotide polymorphism rs1335532 is associated with strong binding of Ascl2 transcription factor and elevated CD58 expression in B-cells.
- Author
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Mitkin NA, Muratova AM, Korneev KV, Pavshintsev VV, Rumyantsev KA, Vagida MS, Uvarova AN, Afanasyeva MA, Schwartz AM, and Kuprash DV
- Subjects
- Alleles, Basic Helix-Loop-Helix Transcription Factors genetics, Binding Sites, CD58 Antigens chemistry, Cell Line, Tumor, Computational Biology methods, Enhancer Elements, Genetic, Female, Gene Expression Regulation, Neoplastic, Genetic Association Studies, Humans, Male, Multiple Sclerosis metabolism, Promoter Regions, Genetic, Wnt Signaling Pathway, Basic Helix-Loop-Helix Transcription Factors metabolism, CD58 Antigens genetics, Multiple Sclerosis genetics, Polymorphism, Single Nucleotide, Up-Regulation
- Abstract
CD58 is expressed on the surface of antigen-presenting cells, including B-cells, and provides co-stimulation to regulatory T-cells (Treg) through CD2 receptor binding. Tregs appear to be essential suppressors of tissue-specific autoimmune responses. Thereby, CD58 plays protective role in multiple sclerosis (MS) and CD58 was identified among several loci associated with MS susceptibility. Minor (C) variant of the single-nucleotide polymorphism (SNP) rs1335532 is associated with lower MS risk according to genome-wide association studies (GWAS) and its presence correlates with higher CD58 mRNA levels in MS patients. We found that genomic region containing rs1335532 has enhancer properties and can significantly boost the CD58 promoter activity in lymphoblast cells. Using bioinformatics and pull-down assay we found that the protective (C) rs1335532 allele created functional binding site for ASCL2 transcription factor, a target of the Wnt signaling pathway. Both in B-lymphoblastoid cell lines and in primary B-cells, as well as in a monocytic cell line, activation of Wnt signaling resulted in an increased CD58 promoter activity in the presence of the protective but not the risk allele of rs1335532, whereas ASCL2 knockdown abrogated this effect. In summary, our results suggest that ASCL2 mediates the protective function of rs1335532 minor (C) allele in MS., (Copyright © 2018 Elsevier B.V. All rights reserved.)
- Published
- 2018
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28. The Risk G Allele of the Single-Nucleotide Polymorphism rs928413 Creates a CREB1-Binding Site That Activates IL33 Promoter in Lung Epithelial Cells.
- Author
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Gorbacheva AM, Korneev KV, Kuprash DV, and Mitkin NA
- Subjects
- Alleles, Asthma metabolism, Cell Line, Tumor, Child, Epithelial Cells metabolism, Genetic Predisposition to Disease, Humans, Lung cytology, Lung metabolism, MAP Kinase Signaling System, Promoter Regions, Genetic, Protein Binding, Respiratory Mucosa cytology, Respiratory Mucosa metabolism, Transcriptional Activation, Asthma genetics, Cyclic AMP Response Element-Binding Protein metabolism, Interleukin-33 genetics, Polymorphism, Single Nucleotide
- Abstract
Cytokine interleukin 33 (IL-33) is constitutively expressed by epithelial barrier cells, and promotes the development of humoral immune responses. Along with other proinflammatory mediators released by the epithelium of airways and lungs, it plays an important role in a number of respiratory pathologies. In particular, IL-33 significantly contributes to pathogenesis of allergy and asthma; genetic variations in the IL33 locus are associated with increased susceptibility to asthma. Large-scale genome-wide association studies have identified minor "G" allele of the single-nucleotide polymorphism rs928413, located in the IL33 promoter area, as a susceptible variant for early childhood and atopic asthma development. Here, we demonstrate that the rs928413(G) allele creates a binding site for the cAMP response element-binding protein 1 (CREB1) transcription factor. In a pulmonary epithelial cell line, activation of CREB1, presumably via the p38 mitogen-activated protein kinases (MAPK) cascade, activates the IL33 promoter containing the rs928413(G) allele specifically and in a CREB1-dependent manner. This mechanism may explain the negative effect of the rs928413 minor "G" allele on asthma development.
- Published
- 2018
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29. [The Novel Short Isoform of Securin Stimulates the Expression of Cyclin D3 and Angiogenesis Factors VEGFA and FGF2, but Does Not Affect the Expression of MYC Transcription Factor].
- Author
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Demin DE, Bogolyubova AV, Zlenko DV, Uvarova AN, Deikin AV, Putlyaeva LV, Belousov PV, Mitkin NA, Korneev KV, Sviryaeva EN, Kulakovskiy IV, Tatosyan KA, Kuprash DV, and Schwartz AM
- Subjects
- Cyclin D3 genetics, Fibroblast Growth Factor 2 genetics, HEK293 Cells, Hep G2 Cells, Humans, Jurkat Cells, K562 Cells, MCF-7 Cells, Protein Isoforms biosynthesis, Protein Isoforms genetics, Proto-Oncogene Proteins c-myc genetics, Securin genetics, Vascular Endothelial Growth Factor A genetics, Cyclin D3 biosynthesis, Fibroblast Growth Factor 2 biosynthesis, Gene Expression Regulation, Neoplastic, Proto-Oncogene Proteins c-myc biosynthesis, Securin metabolism, Vascular Endothelial Growth Factor A biosynthesis
- Abstract
Pituitary tumor-transforming gene-1 (PTTG1) encodes securin, a multifunctional protein involved in development of various types of cancer. Securin participates in the regulation of sister chromatids separation and the expression of multiple genes involved in the control of the cell cycle, metabolism, and angiogenesis. In several human cell lines, we have found a novel short isoform of securin mRNA, which does not contain exons 3 and 4. After the translation of this new mRNA, a shortened protein is produced that, like the full-size form, is able to activate the transcription of cyclin D3 gene (CCND3), which controls the G1/S transition and angiogenesis factors VEGFA (vascular endothelial growth factor), and FGF2 (fibroblast growth factor 2) in HEK293 cells. However, unlike the full-size protein, the short isoform of PTTG1 does not affect the MYC gene expression because it lacks the DNA-binding domain, which is needed for its interactions with the MYC promoter. Furthermore, the short form of securin does not influence the expression of MYC transcriptional targets, such as TP53 and IL-8. Thus, we found a novel isoform of securin which is able to activate a more restricted repertoire of genes compared to the full-size protein.
- Published
- 2018
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30. Hypoacylated LPS from Foodborne Pathogen Campylobacter jejuni Induces Moderate TLR4-Mediated Inflammatory Response in Murine Macrophages.
- Author
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Korneev KV, Kondakova AN, Sviriaeva EN, Mitkin NA, Palmigiano A, Kruglov AA, Telegin GB, Drutskaya MS, Sturiale L, Garozzo D, Nedospasov SA, Knirel YA, and Kuprash DV
- Subjects
- Animals, Campylobacter jejuni pathogenicity, Cytokines metabolism, Interferon Regulatory Factor-3 genetics, Interleukin-1beta metabolism, Interleukin-6, Lipid A immunology, Lipid A isolation & purification, Lipid A pharmacology, Lipopolysaccharides immunology, Macrophages drug effects, Macrophages immunology, Mice, Mice, Inbred C57BL, RNA, Small Interfering, Toll-Like Receptor 4 genetics, Tumor Necrosis Factor-alpha metabolism, Campylobacter jejuni immunology, Campylobacter jejuni metabolism, Foodborne Diseases microbiology, Lipopolysaccharides pharmacology, Toll-Like Receptor 4 drug effects, Toll-Like Receptor 4 immunology
- Abstract
Toll-like receptor 4 (TLR4) initiates immune response against Gram-negative bacteria upon specific recognition of lipid A moiety of lipopolysaccharide (LPS), the major component of their cell wall. Some natural differences between LPS variants in their ability to interact with TLR4 may lead to either insufficient activation that may not prevent bacterial growth, or excessive activation which may lead to septic shock. In this study we evaluated the biological activity of LPS isolated from pathogenic strain of Campylobacter jejuni , the most widespread bacterial cause of foodborne diarrhea in humans. With the help of hydrophobic chromatography and MALDI-TOF mass spectrometry we showed that LPS from a C. jejuni strain O2A consists of both hexaacyl and tetraacyl forms. Since such hypoacylation can result in a reduced immune response in humans, we assessed the activity of LPS from C. jejuni in mouse macrophages by measuring its capacity to activate TLR4-mediated proinflammatory cytokine and chemokine production, as well as NFκB-dependent reporter gene transcription. Our data support the hypothesis that LPS acylation correlates with its bioactivity.
- Published
- 2018
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31. p63 and p73 repress CXCR5 chemokine receptor gene expression in p53-deficient MCF-7 breast cancer cells during genotoxic stress.
- Author
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Mitkin NA, Muratova AM, Sharonov GV, Korneev KV, Sviriaeva EN, Mazurov D, Schwartz AM, and Kuprash DV
- Subjects
- CRISPR-Cas Systems, Female, Humans, MCF-7 Cells, Methyl Methanesulfonate pharmacology, NF-kappa B physiology, Promoter Regions, Genetic, DNA Damage, Gene Expression Regulation, Neoplastic, Membrane Proteins physiology, Receptors, CXCR5 genetics, Tumor Protein p73 physiology, Tumor Suppressor Protein p53 physiology
- Abstract
Many types of chemotherapeutic agents induce of DNA-damage that is accompanied by activation of p53 tumor suppressor, a key regulator of tumor development and progression. In our previous study we demonstrated that p53 could repress CXCR5 chemokine receptor gene in MCF-7 breast cancer cells via attenuation of NFkB activity. In this work we aimed to determine individual roles of p53 family members in the regulation of CXCR5 gene expression under genotoxic stress. DNA-alkylating agent methyl methanesulfonate caused a reduction in CXCR5 expression not only in parental MCF-7 cells but also in MCF-7-p53off cells with CRISPR/Cas9-mediated inactivation of the p53 gene. Since p53 knockout was associated with elevated expression of its p63 and p73 homologues, we knocked out p63 using CRISPR/Cas9 system and knocked down p73 using specific siRNA. The CXCR5 promoter activity, CXCR5 expression and CXCL13-directed migration in MCF-7 cells with inactivation of all three p53 family genes were completely insensitive to genotoxic stress, while pairwise p53+p63 or p53+p73 inactivation resulted in partial effects. Using deletion analysis and site-directed mutagenesis, we demonstrated that effects of NFkB on the CXCR5 promoter inversely correlated with p63 and p73 levels. Thus, all three p53 family members mediate the effects of genotoxic stress on the CXCR5 promoter using the same mechanism associated with attenuation of NFkB activity. Understanding of this mechanism could facilitate prognosis of tumor responses to chemotherapy., (Copyright © 2017 Elsevier B.V. All rights reserved.)
- Published
- 2017
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32. [Antibody-Based Drugs and Other Recombinant Proteins for Diagnostics and Therapy of Viral Infections, Autoimmune Diseases and Cancer].
- Author
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Kuprash DV, Garib FY, and Nedospasov SA
- Subjects
- Animals, Antibodies, Monoclonal biosynthesis, Antibodies, Monoclonal genetics, Antibodies, Neutralizing biosynthesis, Antibodies, Neutralizing genetics, Antibodies, Viral biosynthesis, Antibodies, Viral genetics, Autoimmune Diseases diagnosis, Autoimmune Diseases drug therapy, Autoimmune Diseases pathology, Humans, Neoplasms diagnosis, Neoplasms drug therapy, Neoplasms immunology, Neoplasms pathology, Protein Engineering methods, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins immunology, Virus Diseases diagnosis, Virus Diseases drug therapy, Virus Diseases immunology, Virus Diseases virology, Antibodies, Monoclonal therapeutic use, Antibodies, Neutralizing therapeutic use, Antibodies, Viral therapeutic use, Antineoplastic Agents, Immunological therapeutic use, Recombinant Fusion Proteins therapeutic use
- Published
- 2017
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33. The Minor Variant of the Single-Nucleotide Polymorphism rs3753381 Affects the Activity of a SLAMF1 Enhancer.
- Author
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Putlyaeva LV, Schwartz AM, Klepikova AV, Vorontsov IE, Kulakovskiy IV, and Kuprash DV
- Abstract
The SLAMF1 gene encodes CD150, a transmembrane glycoprotein expressed on the surface of T and B-lymphocytes, NK-cells, dendritic cells, and subpopulations of macrophages and basophils. We investigated the functional regulatory polymorphisms of the SLAMF1 locus associated with autoimmune processes, using bioinformatics and a mutational analysis of the regulatory elements overlapping with polymorphic positions. In the reporter gene assay in MP-1 and Raji B-cell lines, the enhancer activity of the regulatory region of the locus containing the rs3753381 polymorphism demonstrated a twofold increase upon the introduction of the rs3753381 minor variant (G → A) associated with myasthenia gravis. An analysis of the nucleotide context in the vicinity of rs3753381 revealed that the minor version of this polymorphism improves several binding sites for the transcription factors of FOX and NFAT, and RXR nuclear receptors. All mutations that disrupt any of these sites lead to a decrease in the enhancer activity both in MP1 and in Raji cells, and each of the two B-cell lines expresses a specific set of these factors. Thus, the minor variant of the rs3753381 polymorphism may contribute to the development of myasthenia gravis by modulating SLAMF1 expression, presumably in pathogenic B-lymphocytes.
- Published
- 2017
34. Low level of Lck kinase in Th2 cells limits expression of CD4 co-receptor and S73 phosphorylation of transcription factor c-Jun.
- Author
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Shebzukhov YV, Stanislawiak S, Bezhaeva TR, Nedospasov SA, and Kuprash DV
- Subjects
- Animals, Cells, Cultured, Gene Expression Profiling, HEK293 Cells, Humans, Lymphocyte Specific Protein Tyrosine Kinase p56(lck) genetics, Mice, Inbred C57BL, Phosphorylation, Serine metabolism, Signal Transduction, Th1 Cells metabolism, CD4-Positive T-Lymphocytes metabolism, Lymphocyte Specific Protein Tyrosine Kinase p56(lck) metabolism, Proto-Oncogene Proteins c-jun metabolism, Receptors, Antigen, T-Cell metabolism, Th2 Cells metabolism
- Abstract
The Src-family tyrosine kinase Lck is an enzyme associated with the CD4 and CD8 co-receptors and promoting signaling through the T cell receptor (TCR) complex. The levels of Lck expression and activity change during the development and differentiation of T cells. Here we show that Lck expression is higher in Th1 cells as compared to Th2 cells. Ectopic overexpression of Lck in Th2 cells results in increased expression of CD4 co-receptor and enhanced S73 phosphorylation of transcription factor c-Jun. Our findings indicate that TCR-mediated signaling in Th2 cells may be directly attenuated by Lck protein expression level.
- Published
- 2017
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35. The single nucleotide variant rs12722489 determines differential estrogen receptor binding and enhancer properties of an IL2RA intronic region.
- Author
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Afanasyeva MA, Putlyaeva LV, Demin DE, Kulakovskiy IV, Vorontsov IE, Fridman MV, Makeev VJ, Kuprash DV, and Schwartz AM
- Subjects
- Alleles, Base Sequence, Binding Sites, Cell Line, Transformed, Chromatin Immunoprecipitation, Electrophoretic Mobility Shift Assay, Estrogen Receptor alpha metabolism, Gene Expression Regulation, Genes, Reporter, Human T-lymphotropic virus 1 genetics, Humans, Interleukin-2 Receptor alpha Subunit metabolism, Introns, Luciferases genetics, Luciferases metabolism, Protein Binding, T-Lymphocytes, Regulatory cytology, Estrogen Receptor alpha genetics, Interleukin-2 Receptor alpha Subunit genetics, Polymorphism, Single Nucleotide, Response Elements, T-Lymphocytes, Regulatory metabolism
- Abstract
We studied functional effect of rs12722489 single nucleotide polymorphism located in the first intron of human IL2RA gene on transcriptional regulation. This polymorphism is associated with multiple autoimmune conditions (rheumatoid arthritis, multiple sclerosis, Crohn's disease, and ulcerative colitis). Analysis in silico suggested significant difference in the affinity of estrogen receptor (ER) binding site between alternative allelic variants, with stronger predicted affinity for the risk (G) allele. Electrophoretic mobility shift assay showed that purified human ERα bound only G variant of a 32-bp genomic sequence containing rs12722489. Chromatin immunoprecipitation demonstrated that endogenous human ERα interacted with rs12722489 genomic region in vivo and DNA pull-down assay confirmed differential allelic binding of amplified 189-bp genomic fragments containing rs12722489 with endogenous human ERα. In a luciferase reporter assay, a kilobase-long genomic segment containing G but not A allele of rs12722489 demonstrated enhancer properties in MT-2 cell line, an HTLV-1 transformed human cell line with a regulatory T cell phenotype.
- Published
- 2017
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36. Multiple single nucleotide polymorphisms in the first intron of the IL2RA gene affect transcription factor binding and enhancer activity.
- Author
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Schwartz AM, Demin DE, Vorontsov IE, Kasyanov AS, Putlyaeva LV, Tatosyan KA, Kulakovskiy IV, and Kuprash DV
- Subjects
- Autoimmune Diseases genetics, Autoimmune Diseases immunology, Cell Line, Enhancer Elements, Genetic, Genetic Predisposition to Disease, Humans, Introns, Jurkat Cells, Polymorphism, Single Nucleotide, Promoter Regions, Genetic, T-Lymphocytes, Helper-Inducer immunology, T-Lymphocytes, Helper-Inducer metabolism, T-Lymphocytes, Regulatory immunology, T-Lymphocytes, Regulatory metabolism, Transcription Factors metabolism, Interleukin-2 Receptor alpha Subunit genetics
- Abstract
IL2RA gene encodes the alpha subunit of a high-affinity receptor for interleukin-2 which is expressed by several distinct populations of lymphocytes involved in autoimmune processes. A large number of polymorphic alleles of the IL2RA locus are associated with the development of various autoimmune diseases. With bioinformatics analysis we the dissected the first intron of the IL2RA gene and selected several single nucleotide polymorphisms (SNPs) that may influence the regulation of the IL2RA gene in cell types relevant to autoimmune pathology. We described five enhancers containing the selected SNPs that stimulated activity of the IL2RA promoter in a cell-type specific manner, and tested the effect of specific SNP alleles on activity of the respective enhancers (E1 to E5, labeled according to the distance to the promoter). The E4 enhancer with minor T variant of rs61839660 SNP demonstrated reduced activity due to disrupted binding of MEF2A/C transcription factors (TFs). Neither rs706778 nor rs706779 SNPs, both associated with a number of autoimmune diseases, had any effect on the activity of the enhancer E2. However, rare variants of several SNPs (rs139767239, rs115133228, rs12722502, rs12722635) genetically linked to either rs706778 and/or rs706779 significantly influenced the activity of E1, E3 and E5 enhancers, presumably by disrupting EBF1, GABPA and ELF1 binding sites., (Copyright © 2016 Elsevier B.V. All rights reserved.)
- Published
- 2017
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37. TLR-signaling and proinflammatory cytokines as drivers of tumorigenesis.
- Author
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Korneev KV, Atretkhany KN, Drutskaya MS, Grivennikov SI, Kuprash DV, and Nedospasov SA
- Subjects
- Animals, Cell Transformation, Neoplastic pathology, Cytokines immunology, Humans, Neoplasms pathology, Cell Transformation, Neoplastic immunology, Neoplasm Proteins immunology, Neoplasms immunology, Signal Transduction immunology, Toll-Like Receptor 4 immunology, Tumor Microenvironment immunology
- Abstract
The link between inflammation and cancer was first proposed by R. Virchow. It was later realized that it is chronic inflammation that may promote cancer, whereas acute inflammation can actually block tumor development or even result in cure. Many molecular mediators of these diverse processes have been characterized only during the past 3 decades thanks to the advances in molecular and cellular techniques, as well as due to technologies of reverse genetics. In this chapter we discuss the role of Toll-like receptor (TLR) 4 signaling in cancer and contributions of proinflammatory cytokine signaling (whose expression may be driven by TLR-mediated signals) to tumor-promoting microenvironment. We also discuss recent clinical advances to target these pro-tumorigenic pathways at distinct stages of tumorigenesis., (Copyright © 2016 Elsevier Ltd. All rights reserved.)
- Published
- 2017
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38. Chemokines, cytokines and exosomes help tumors to shape inflammatory microenvironment.
- Author
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Atretkhany KN, Drutskaya MS, Nedospasov SA, Grivennikov SI, and Kuprash DV
- Subjects
- Animals, Chemokines metabolism, Cytokines metabolism, Disease Progression, Exosomes metabolism, Humans, Inflammation immunology, Neoplasms immunology, Inflammation pathology, Neoplasms pathology, Tumor Microenvironment immunology
- Abstract
Relationship between inflammation and cancer is now well-established and represents a paradigm that our immune response does not necessarily serves solely to protect us from infections and cancer. Many specific mechanisms that link chronic inflammation to cancer promotion and metastasis have been uncovered in the recent years. Here we are focusing on the effects that tumors may exert on inflammatory cascades, tuning the immune system ability to cause tumor promotion or regression. In particular, we discuss the contributions of chemokines, cytokines and exosomes to the processes such as induction of inflammation and tumorigenesis. Overall, tumor-elicited inflammation is a key driver of tumor progression and an essential component of tumor microenvironment., (Copyright © 2016 Elsevier Inc. All rights reserved.)
- Published
- 2016
- Full Text
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39. The A Allele of the Single-Nucleotide Polymorphism rs630923 Creates a Binding Site for MEF2C Resulting in Reduced CXCR5 Promoter Activity in B-Cell Lymphoblastic Cell Lines.
- Author
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Mitkin NA, Muratova AM, Schwartz AM, and Kuprash DV
- Abstract
Chemokine receptor CXCR5 is highly expressed in B-cells and under normal conditions is involved in their migration to specific areas of secondary lymphoid organs. B-cells are known to play an important role in various autoimmune diseases including multiple sclerosis (MS) where areas of demyelinating lesions attract B-cells by overexpressing CXCL13, the CXCR5 ligand. In this study, we aimed to determine the functional significance of single-nucleotide polymorphism rs630923 (A/C), which is located in cxcr5 gene promoter, and its common allele is associated with increased risk of MS. Using bioinformatics and pull-down assay in B-lymphoblastic cell lines, we showed that protective minor rs630923 "A" allele created functional binding site for MEF2C transcription factor. Elevated MEF2C expression in B-cells correlated with reduced activity of cxcr5 promoter containing rs630923 "A" allele. This effect that was fully neutralized by MEF2C-directed siRNA may mechanistically explain the protective role of the rs630923 minor allele in MS. Using site-directed mutagenesis of the cxcr5 gene promoter, we were unable to find any experimental evidence for the previously proposed role of NFκB transcription factors in rs630923-mediated CXCR5 promoter regulation. Thus, our results identify MEF2C as a possible mediator of protective function of the rs630923 "A" allele in MS.
- Published
- 2016
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40. Molecular and Cellular Mechanisms of Inflammation.
- Author
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Kuprash DV and Nedospasov SA
- Subjects
- Animals, Humans, Inflammation pathology, Inflammation genetics, Inflammation immunology
- Abstract
Inflammation is one of the most fundamental and pronounced protective reactions of the organism. From ancient times to the present day, complex and diverse patterns of inflammation development and their role in various diseases have attracted attention of investigators. This issue of Biokhimiya/Biochemistry (Moscow) includes experimental studies and reviews dedicated to various aspects of this important and interesting problem.
- Published
- 2016
- Full Text
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41. Modeling of viral-bacterial coinfections at the molecular level using agonists of innate immunity receptors.
- Author
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Sviriaeva EN, Korneev KV, Drutskaya MS, Nedospasov SA, and Kuprash DV
- Subjects
- Aminoquinolines pharmacology, Animals, Bacterial Infections immunology, Bacterial Infections metabolism, Bone Marrow, Cells, Cultured, Coinfection immunology, Coinfection metabolism, Disease Models, Animal, Imiquimod, Immunity, Innate, Interleukin-1beta genetics, Interleukin-1beta metabolism, Interleukin-6 genetics, Interleukin-6 metabolism, Ligands, Lipopolysaccharides immunology, Membrane Glycoproteins agonists, Membrane Glycoproteins metabolism, Mice, Mice, Inbred C57BL, RNA, Messenger metabolism, Toll-Like Receptor 4 agonists, Toll-Like Receptor 4 metabolism, Toll-Like Receptor 7 agonists, Toll-Like Receptor 7 metabolism, Tumor Necrosis Factor-alpha genetics, Tumor Necrosis Factor-alpha metabolism, Virus Diseases immunology, Virus Diseases metabolism, Cytokines genetics, Macrophage Activation, Macrophages immunology, Membrane Glycoproteins immunology, Toll-Like Receptor 4 immunology, Toll-Like Receptor 7 immunology, Toll-Like Receptors agonists, Toll-Like Receptors metabolism
- Abstract
The combined effect of innate immunity receptors in viral-bacterial coinfections was studied in vitro using the primary culture of murine macrophages activated by different combinations of ligands of innate immunity receptors belonging to the family of Toll-like receptors. The activation of macrophages first with a viral ligand and then with a bacterial one significantly decreased the expression of proinflammatory cytokine genes. Such attenuation of immune responses may occur during the development of bacterial complications in viral infections.
- Published
- 2016
- Full Text
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42. Mechanisms of Changes in Immune Response during Bacterial Coinfections of the Respiratory Tract.
- Author
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Sviriaeva EN, Korneev KV, Drutskaya MS, and Kuprash DV
- Subjects
- Animals, Bacterial Infections pathology, Humans, Lung pathology, Respiratory Tract Infections pathology, Bacterial Infections immunology, Lung immunology, Respiratory Tract Infections immunology
- Abstract
Acute diseases of the respiratory tract are often caused by viral pathogens and accompanying secondary bacterial infections. It is known that the development of such bacterial complications is caused mainly by a decreased infiltration with immune system cells and by suppressed inflammation in the lungs. There are significant advances in understanding the mechanisms of secondary infections, although many details remain unclear. This review summarizes current knowledge of the molecular and cellular changes in the host organism that can influence the course of bacterial coinfections in the respiratory tract.
- Published
- 2016
- Full Text
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43. Early B-cell factor 1 (EBF1) is critical for transcriptional control of SLAMF1 gene in human B cells.
- Author
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Schwartz AM, Putlyaeva LV, Covich M, Klepikova AV, Akulich KA, Vorontsov IE, Korneev KV, Dmitriev SE, Polanovsky OL, Sidorenko SP, Kulakovskiy IV, and Kuprash DV
- Subjects
- B-Lymphocytes cytology, B-Lymphocytes metabolism, Basic Helix-Loop-Helix Transcription Factors genetics, Basic Helix-Loop-Helix Transcription Factors metabolism, Binding Sites, Cell Line, Tumor, Enhancer Elements, Genetic, Genes, Reporter, HEK293 Cells, Humans, Interferon Regulatory Factors genetics, Interferon Regulatory Factors metabolism, Luciferases genetics, Mutation, NF-kappa B genetics, NF-kappa B metabolism, Nuclear Proteins genetics, Nuclear Proteins metabolism, Primary Cell Culture, Promoter Regions, Genetic, Protein Binding, Proto-Oncogene Proteins genetics, Proto-Oncogene Proteins metabolism, STAT6 Transcription Factor genetics, STAT6 Transcription Factor metabolism, Signal Transduction, Signaling Lymphocytic Activation Molecule Family Member 1 metabolism, Sp1 Transcription Factor genetics, Sp1 Transcription Factor metabolism, Trans-Activators metabolism, Transcription Factors genetics, Transcription Factors metabolism, Gene Expression Regulation, Signaling Lymphocytic Activation Molecule Family Member 1 genetics, Trans-Activators genetics, Transcription, Genetic
- Abstract
Signaling lymphocytic activation molecule family member 1 (SLAMF1)/CD150 is a co-stimulatory receptor expressed on a variety of hematopoietic cells, in particular on mature lymphocytes activated by specific antigen, costimulation and cytokines. Changes in CD150 expression level have been reported in association with autoimmunity and with B-cell chronic lymphocytic leukemia. We characterized the core promoter for SLAMF1 gene in human B-cell lines and explored binding sites for a number of transcription factors involved in B cell differentiation and activation. Mutations of SP1, STAT6, IRF4, NF-kB, ELF1, TCF3, and SPI1/PU.1 sites resulted in significantly decreased promoter activity of varying magnitude, depending on the cell line tested. The most profound effect on the promoter strength was observed upon mutation of the binding site for Early B-cell factor 1 (EBF1). This mutation produced a 10-20 fold drop in promoter activity and pinpointed EBF1 as the master regulator of human SLAMF1 gene in B cells. We also identified three potent transcriptional enhancers in human SLAMF1 locus, each containing functional EBF1 binding sites. Thus, EBF1 interacts with specific binding sites located both in the promoter and in the enhancer regions of the SLAMF1 gene and is critical for its expression in human B cells., (Copyright © 2016 Elsevier B.V. All rights reserved.)
- Published
- 2016
- Full Text
- View/download PDF
44. [Genetic mechanisms of adaptive immunity emergence in vertebrates].
- Author
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Shilov ES and Kuprash DV
- Subjects
- Animals, Antibodies immunology, Genetic Loci immunology, Humans, Receptors, Antigen, T-Cell immunology, Adaptive Immunity genetics, Antibodies genetics, Antigen Presentation genetics, Major Histocompatibility Complex, Receptors, Antigen, T-Cell genetics, Transplantation Immunology genetics
- Abstract
The adaptive immune system in vertebrates emerged in a multistep process that can be reconstructed on the basis of the data concerning the structure of immune systems of modern cartilaginous and bony fishes, as well as of cyclostomes. The most probable evolutionary scenario is likely to be as follows: the T cell receptor loci emerged on the basis of NK cell-like receptor genes; the antibody loci evolved on the basis of T cell receptor loci; the MHC locus arose on the basis of the locus responsible for innate immunity of early chordates. The ancestral MHC molecules likely participated in the transplantation immunity before they acquired the ability of antigen peptide presentation.
- Published
- 2016
45. Erratum to: CXCL13-CXCR5 co-expression regulates epithelial to mesenchymal transition of breast cancer cells during lymph node metastasis.
- Author
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Biswas S, Sengupta S, Roy Chowdhury S, Jana S, Mandal G, Mandal PK, Saha N, Malhotra V, Gupta A, Kuprash DV, and Bhattacharyya A
- Published
- 2016
- Full Text
- View/download PDF
46. Structural Relationship of the Lipid A Acyl Groups to Activation of Murine Toll-Like Receptor 4 by Lipopolysaccharides from Pathogenic Strains of Burkholderia mallei, Acinetobacter baumannii, and Pseudomonas aeruginosa.
- Author
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Korneev KV, Arbatsky NP, Molinaro A, Palmigiano A, Shaikhutdinova RZ, Shneider MM, Pier GB, Kondakova AN, Sviriaeva EN, Sturiale L, Garozzo D, Kruglov AA, Nedospasov SA, Drutskaya MS, Knirel YA, and Kuprash DV
- Abstract
Toll-like receptor 4 (TLR4) is required for activation of innate immunity upon recognition of lipopolysaccharide (LPS) of Gram-negative bacteria. The ability of TLR4 to respond to a particular LPS species is important since insufficient activation may not prevent bacterial growth while excessive immune reaction may lead to immunopathology associated with sepsis. Here, we investigated the biological activity of LPS from Burkholderia mallei that causes glanders, and from the two well-known opportunistic pathogens Acinetobacter baumannii and Pseudomonas aeruginosa (causative agents of nosocomial infections). For each bacterial strain, R-form LPS preparations were purified by hydrophobic chromatography and the chemical structure of lipid A, an LPS structural component, was elucidated by HR-MALDI-TOF mass spectrometry. The biological activity of LPS samples was evaluated by their ability to induce production of proinflammatory cytokines, such as IL-6 and TNF, by bone marrow-derived macrophages. Our results demonstrate direct correlation between the biological activity of LPS from these pathogenic bacteria and the extent of their lipid A acylation.
- Published
- 2015
- Full Text
- View/download PDF
47. Control of Mycobacterial Infections in Mice Expressing Human Tumor Necrosis Factor (TNF) but Not Mouse TNF.
- Author
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Olleros ML, Chavez-Galan L, Segueni N, Bourigault ML, Vesin D, Kruglov AA, Drutskaya MS, Bisig R, Ehlers S, Aly S, Walter K, Kuprash DV, Chouchkova M, Kozlov SV, Erard F, Ryffel B, Quesniaux VF, Nedospasov SA, and Garcia I
- Subjects
- Animals, Blotting, Western, Cytokines biosynthesis, Flow Cytometry, Gene Knock-In Techniques methods, Humans, Macrophages immunology, Mice, Mice, Inbred C57BL, Mice, Knockout, Mycobacterium bovis immunology, Mycobacterium tuberculosis immunology, Disease Models, Animal, Mycobacterium Infections immunology, Tumor Necrosis Factor-alpha genetics, Tumor Necrosis Factor-alpha immunology
- Abstract
Tumor necrosis factor (TNF) is an important cytokine for host defense against pathogens but is also associated with the development of human immunopathologies. TNF blockade effectively ameliorates many chronic inflammatory conditions but compromises host immunity to tuberculosis. The search for novel, more specific human TNF blockers requires the development of a reliable animal model. We used a novel mouse model with complete replacement of the mouse TNF gene by its human ortholog (human TNF [huTNF] knock-in [KI] mice) to determine resistance to Mycobacterium bovis BCG and M. tuberculosis infections and to investigate whether TNF inhibitors in clinical use reduce host immunity. Our results show that macrophages from huTNF KI mice responded to BCG and lipopolysaccharide similarly to wild-type macrophages by NF-κB activation and cytokine production. While TNF-deficient mice rapidly succumbed to mycobacterial infection, huTNF KI mice survived, controlling the bacterial burden and activating bactericidal mechanisms. Administration of TNF-neutralizing biologics disrupted the control of mycobacterial infection in huTNF KI mice, leading to an increased bacterial burden and hyperinflammation. Thus, our findings demonstrate that human TNF can functionally replace murine TNF in vivo, providing mycobacterial resistance that could be compromised by TNF neutralization. This new animal model will be helpful for the testing of specific biologics neutralizing human TNF., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)
- Published
- 2015
- Full Text
- View/download PDF
48. Serum Immunoproteomics Combined With Pathological Reassessment of Surgical Specimens Identifies TCP-1ζ Autoantibody as a Potential Biomarker in Thyroid Neoplasia.
- Author
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Belousov PV, Bogolyubova AV, Kim YS, Abrosimov AY, Kopylov AT, Tvardovskiy AA, Lanshchakov KV, Sazykin AY, Dvinskikh NY, Bobrovskaya YI, Selivanova LS, Shilov ES, Schwartz AM, Shebzukhov YV, Severskaia NV, Vanushko VE, Moshkovskii SA, Nedospasov SA, and Kuprash DV
- Subjects
- Adenocarcinoma, Follicular blood, Adenocarcinoma, Follicular immunology, Adenocarcinoma, Follicular pathology, Adult, Biomarkers blood, Carcinoma, Papillary blood, Carcinoma, Papillary immunology, Carcinoma, Papillary pathology, Female, Humans, Middle Aged, Thyroid Neoplasms blood, Thyroid Neoplasms immunology, Thyroid Neoplasms pathology, Young Adult, Adenocarcinoma, Follicular diagnosis, Autoantibodies blood, Carcinoma, Papillary diagnosis, Chaperonin Containing TCP-1 immunology, Thyroid Neoplasms diagnosis
- Abstract
Context: Current methods of preoperative diagnostics frequently fail to discriminate between benign and malignant thyroid neoplasms. In encapsulated follicular-patterned tumors (EnFPT), this discrimination is challenging even using histopathological analysis. Autoantibody response against tumor-associated antigens is a well-documented phenomenon with prominent diagnostic potential; however, autoantigenicity of thyroid tumors remains poorly explored., Objectives: Objectives were exploration of tumor-associated antigen repertoire of thyroid tumors and identification of candidate autoantibody biomarkers capable of discrimination between benign and malignant thyroid neoplasms., Design, Setting, and Patients: Proteins isolated from FTC-133 cells were subjected to two-dimensional Western blotting using pooled serum samples of patients originally diagnosed with either papillary thyroid carcinoma (PTC) or EnFPT represented by apparently benign follicular thyroid adenomas, as well as healthy individuals. Immunoreactive proteins were identified using liquid chromatography-tandem mass-spectrometry. Pathological reassessment of EnFPT was performed applying nonconservative criteria for capsular invasion and significance of focal PTC nuclear changes (PTC-NCs). Recombinant T-complex protein 1 subunitζ (TCP-1ζ) was used to examine an expanded serum sample set of patients with various thyroid neoplasms (n = 89) for TCP-1ζ autoantibodies. All patients were included in tertiary referral centers., Results: A protein demonstrating a distinct pattern of EnFPT-specific seroreactivity was identified as TCP-1ζ protein. A subsequent search for clinicopathological correlates of TCP-1ζ seroreactivity revealed nonclassical capsular invasion or focal PTC-NC in all TCP-1ζ antibody-positive cases. Further studies in an expanded sample set confirmed the specificity of TCP-1ζ autoantibodies to malignant EnFPT., Conclusions: We identified TCP-1ζ autoantibodies as a potential biomarker for presurgical discrimination between benign and malignant encapsulated follicular-patterned thyroid tumors. Our results suggest the use of nonconservative morphological criteria for diagnosis of malignant EnFPT in biomarker identification studies and provide a peculiar example of uncovering the diagnostic potential of a candidate biomarker using incorporation of pathological reassessment in the pipeline of immunoproteomic research.
- Published
- 2015
- Full Text
- View/download PDF
49. p53-dependent expression of CXCR5 chemokine receptor in MCF-7 breast cancer cells.
- Author
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Mitkin NA, Hook CD, Schwartz AM, Biswas S, Kochetkov DV, Muratova AM, Afanasyeva MA, Kravchenko JE, Bhattacharyya A, and Kuprash DV
- Subjects
- Chemokine CXCL13 metabolism, Chemotaxis genetics, Chemotaxis immunology, Computational Biology, Enhancer Elements, Genetic, Female, Gene Knockdown Techniques, Humans, MCF-7 Cells, NF-kappa B metabolism, Promoter Regions, Genetic, Protein Binding, Receptors, CXCR5 metabolism, Response Elements, Signal Transduction, Transcriptional Activation, Tumor Suppressor Protein p53 genetics, Breast Neoplasms genetics, Breast Neoplasms metabolism, Gene Expression Regulation, Neoplastic, Receptors, CXCR5 genetics, Tumor Suppressor Protein p53 metabolism
- Abstract
Elevated expression of chemokine receptors in tumors has been reported in many instances and is related to a number of survival advantages for tumor cells including abnormal activation of prosurvival intracellular pathways. In this work we demonstrated an inverse correlation between expression levels of p53 tumor suppressor and CXCR5 chemokine receptor in MCF-7 human breast cancer cell line. Lentiviral transduction of MCF-7 cells with p53 shRNA led to elevated CXCR5 at both mRNA and protein levels. Functional activity of CXCR5 in p53-knockdown MCF-7 cells was also increased as shown by activation of target gene expression and chemotaxis in response to B-lymphocyte chemoattractant CXCL13. Using deletion analysis and site-directed mutagenesis of the cxcr5 gene promoter and enhancer elements, we demonstrated that p53 appears to act upon cxcr5 promoter indirectly, by repressing the activity of NFκB transcription factors. Using chromatin immunoprecipitation and reporter gene analysis, we further demonstrated that p65/RelA was able to bind the cxcr5 promoter in p53-dependent manner and to directly transactivate it when overexpressed. Through the described mechanism, elevated CXCR5 expression may contribute to abnormal cell survival and migration in breast tumors that lack functional p53.
- Published
- 2015
- Full Text
- View/download PDF
50. Upstream open reading frames regulate translation of the long isoform of SLAMF1 mRNA that encodes costimulatory receptor CD150.
- Author
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Putlyaeva LV, Schwartz AM, Korneev KV, Covic M, Uroshlev LA, Makeev VY, Dmitriev SE, and Kuprash DV
- Subjects
- 5' Untranslated Regions genetics, Eukaryotic Initiation Factor-2 deficiency, Eukaryotic Initiation Factor-4E deficiency, Genes, Reporter genetics, HEK293 Cells, Humans, Mutagenesis, Site-Directed, Signaling Lymphocytic Activation Molecule Family Member 1, Antigens, CD biosynthesis, Antigens, CD genetics, Open Reading Frames genetics, Protein Biosynthesis genetics, RNA Isoforms genetics, Receptors, Cell Surface biosynthesis, Receptors, Cell Surface genetics
- Abstract
More than 40% of human genes contain upstream open reading frames (uORF) in their 5'-untranslated regions (5'-UTRs) and at the same time express at least one truncated mRNA isoform containing no uORF. We studied translational regulation by four uORFs found in the 5'-UTR of full-length mRNA for SLAMF1, the gene encoding CD150 membrane protein. CD150 is a member of the CD2 superfamily, a costimulatory lymphocyte receptor, a receptor for measles virus, and a microbial sensor on macrophages. The SLAMF1 gene produces at least two mRNA isoforms that differ in their 5'-UTRs. In the long isoform of the SLAMF1 mRNA that harbors four uORFs in the 5'-UTR, the stop codon of uORF4 overlaps with the AUG codon of the main ORF forming a potential termination-reinitiation site UGAUG, while uORF2 and uORF3 start codons flank a sequence identical to Motif 1 from the TURBS regulatory element. TURBS was shown to be required for a coupled termination-reinitiation event during translation of polycistronic RNAs of some viruses. In a model cell system, reporter mRNA based on the 5'-UTR of SLAMF1 short isoform, which lacks any uORF, is translated 5-6 times more efficiently than the mRNA with 5'-UTR from the long isoform. Nucleotide substitutions disrupting start codons in either uORF2-4 result in significant increase in translation efficiency, while substitution of two nucleotides in TURBS Motif 1 leads to a 2-fold decrease in activity. These data suggest that TURBS-like elements can serve for translation control of certain cellular mRNAs containing uORFs.
- Published
- 2014
- Full Text
- View/download PDF
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