Construction and Functional Analysis of Luciferase Reporter Plasmid Containing CD25 Gene Promoter
American Journal of Life Sciences
Volume 5, Issue 6, December 2017, Pages: 160-163
Received: Dec. 27, 2017;
Published: Dec. 28, 2017
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Yuan Xiang, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
Mei Wu, Department of Clinical Microbiology, School of Laboratory Medicine, Hubei University of Chinese Medicine, Wuhan, P.R.China
Jia-Peng Li, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
Zi-Jian Zhang, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
Zhou-Tong Dai, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
Feng Huang, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
Han-Han Li, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
Xing-Hua Liao, Department of Molecular Biology, Institute of Biology and Medicine, College of Life and Health Sciences, Wuhan University of Science and Technology, Wuhan, P.R.China
CD25 is the alpha chain of IL-2R and IL-2R is distributed on the surface of activated T cells, B cells, and NK cell. The transcriptional factor CD25 plays an important role in the proliferation and differentiation of Treg cells. MKL-1 is also called MAL (megakaryocytic acute leukemia) and belongs to the Myocardin family which also includes MKL-2 (megakaryoblastic leukemia 2) and Myocardin. Studies have shown that in the process of mouse megakaryocyte differentiation, the expression of MKL-1 is up-regulated, and MKL-1 promotes the physiological maturation of human and mouse megakaryocytes. However, the exact mechanism of the CD25 on megakaryocytes is unclear. In this study, human CD25 promoter luciferase reporter construct was generated by PCR amplification. Then the PCR fragment was digested and cloned into pGL3 vector, and this promoter sequence was verified by sequencing. The result showed that luciferase reporter with CD25 promoter was successfully constructed. The expression vectors of human MKL-1 were bought from addgene. Then the activation of the CD25 promoter was detected in 293T cells by luciferase reporter assay after transfected expression vectors of human MKL-1. The result showed that transfected the expression vectors can enhance the transcriptional activity of CD25. Our research will reveal the effect of CD25 on megakaryocytes and may provide a theoretical basis and therapeutic method for diseases caused by megakaryocytes.
Construction and Functional Analysis of Luciferase Reporter Plasmid Containing CD25 Gene Promoter, American Journal of Life Sciences.
Vol. 5, No. 6,
2017, pp. 160-163.
T. Nishimoto, M. Kuwana, CD4+CD25+Foxp3+ regulatory T cells in the pathophysiology of immune thrombocytopenia, Seminars in hematology, 50 Suppl 1 (2013) S43-49.
R. Kapur, M. D. Catalina, R. Aslam, E. R. Speck, R. F. Francovitch, J. W. Semple, A highly purified form of staphylococcal protein A alleviates murine immune thrombocytopenia (ITP), British journal of haematology, DOI 10.1111/bjh.14985(2017).
S. Borst, X. Sim, M. Poncz, D. L. French, P. Gadue, Induced Pluripotent Stem Cell-Derived Megakaryocytes and Platelets for Disease Modeling and Clinical Use, Arteriosclerosis, thrombosis, and vascular biology, DOI 10.1161/ATVBAHA.117.309197(2017).
K. Hayashi, T. Morita, Differences in the nuclear export mechanism between myocardin and myocardin-related transcription factor A, The Journal of biological chemistry, 288 (2013) 5743-5755.
G. Huet, E. K. Rajakyla, T. Viita, K. P. Skarp, M. Crivaro, J. Dopie, M. K. Vartiainen, Actin-regulated feedback loop based on Phactr4, PP1 and cofilin maintains the actin monomer pool, Journal of cell science, 126 (2013) 497-507.
K. L. Du, M. Chen, J. Li, J. J. Lepore, P. Mericko, M. S. Parmacek, Megakaryoblastic leukemia factor-1 transduces cytoskeletal signals and induces smooth muscle cell differentiation from undifferentiated embryonic stem cells, The Journal of biological chemistry, 279 (2004) 17578-17586.
Z. Han, X. Li, J. Wu, E. N. Olson, A myocardin-related transcription factor regulates activity of serum response factor in Drosophila, Proceedings of the National Academy of Sciences of the United States of America, 101 (2004) 12567-12572.
T. Sasazuki, T. Sawada, S. Sakon, T. Kitamura, T. Kishi, T. Okazaki, M. Katano, M. Tanaka, M. Watanabe, H. Yagita, K. Okumura, H. Nakano, Identification of a novel transcriptional activator, BSAC, by a functional cloning to inhibit tumor necrosis factor-induced cell death, The Journal of biological chemistry, 277 (2002) 28853-28860.
T. Morita, T. Mayanagi, K. Sobue, Dual roles of myocardin-related transcription factors in epithelial mesenchymal transition via slug induction and actin remodeling, The Journal of cell biology, 179 (2007) 1027-1042.
E. C. Cheng, Q. Luo, E. M. Bruscia, M. J. Renda, J. A. Troy, S. A. Massaro, D. Tuck, V. Schulz, S. M. Mane, N. Berliner, Y. Sun, S. W. Morris, C. Qiu, D. S. Krause, Role for MKL1 in megakaryocytic maturation, Blood, 113 (2009) 2826-2834.
N. Polentarutti, P. Allavena, G. Bianchi, G. Giardina, A. Basile, S. Sozzani, A. Mantovani, M. Introna, IL-2-regulated expression of the monocyte chemotactic protein-1 receptor (CCR2) in human NK cells: characterization of a predominant 3.4-kilobase transcript containing CCR2B and CCR2A sequences, J Immunol, 158 (1997) 2689-2694.
J. W. Leong, J. M. Chase, R. Romee, S. E. Schneider, R. P. Sullivan, M. A. Cooper, T. A. Fehniger, Preactivation with IL-12, IL-15, and IL-18 induces CD25 and a functional high-affinity IL-2 receptor on human cytokine-induced memory-like natural killer cells, Biology of blood and marrow transplantation: journal of the American Society for Blood and Marrow Transplantation, 20 (2014) 463-473.
H. Li, J. Ge, H. Zhao, W. Du, J. Xu, T. Sui, L. Ma, Z. Zhou, A. Qi, R. Yang, Association of cytotoxic T-lymphocyte antigen 4 gene polymorphisms with idiopathic thrombocytopenic purpura in a Chinese population, Platelets, 22 (2011) 39-44.
B. Liu, H. Zhao, M. C. Poon, Z. Han, D. Gu, M. Xu, H. Jia, R. Yang, Z. C. Han, Abnormality of CD4(+)CD25(+) regulatory T cells in idiopathic thrombocytopenic purpura, European journal of haematology, 78 (2007) 139-143.
R. McMillan, The pathogenesis of chronic immune (idiopathic) thrombocytopenic purpura, Seminars in hematology, 37 (2000) 5-9.
A. M. Thornton, E. M. Shevach, CD4+CD25+ immunoregulatory T cells suppress polyclonal T cell activation in vitro by inhibiting interleukin 2 production, The Journal of experimental medicine, 188 (1998) 287-296.
E. M. Shevach, Regulatory T cells in autoimmmunity*, Annual review of immunology, 18 (2000) 423-449.
S. Sakaguchi, Naturally arising CD4+ regulatory t cells for immunologic self-tolerance and negative control of immune responses, Annual review of immunology, 22 (2004) 531-562.
J. M. Miano, Serum response factor: toggling between disparate programs of gene expression, Journal of molecular and cellular cardiology, 35 (2003) 577-593.
G. K. Owens, Molecular control of vascular smooth muscle cell differentiation and phenotypic plasticity, Novartis Foundation symposium, 283 (2007) 174-191; discussion 191-173, 238-141.
Y. Nagata, M. R. Jones, H. G. Nguyen, D. J. McCrann, C. St Hilaire, B. M. Schreiber, A. Hashimoto, M. Inagaki, W. C. Earnshaw, K. Todokoro, K. Ravid, Vascular smooth muscle cell polyploidization involves changes in chromosome passenger proteins and an endomitotic cell cycle, Experimental cell research, 305 (2005) 277-291.
L. R. Penke, S. K. Huang, E. S. White, M. Peters-Golden, Prostaglandin E2 inhibits alpha-smooth muscle actin transcription during myofibroblast differentiation via distinct mechanisms of modulation of serum response factor and myocardin-related transcription factor-A, The Journal of biological chemistry, 289 (2014) 17151-17162.