1.Heme-binding-mediated negative regulation of the tryptophan metabolic enzyme indoleamine 2,3-dioxygenase 1 (IDO1) by IDO2.
Young Kwan LEE ; Hoon Bok LEE ; Dong Mi SHIN ; Min Jueng KANG ; Eugene C YI ; Seungjoo NOH ; Jaewoo LEE ; Chulbom LEE ; Chang Ki MIN ; Eun Young CHOI
Experimental & Molecular Medicine 2014;46(11):e121-
Indoleamine 2,3-dioxygenases (IDOs) are tryptophan-catabolizing enzymes with immunomodulatory functions. However, the biological role of IDO2 and its relationship with IDO1 are unknown. To assess the relationship between IDO2 and IDO1, we investigated the effects of co-expression of human (h) IDO2 on hIDO1 activity. Cells co-expressing hIDO1 and hIDO2 showed reduced tryptophan metabolic activity compared with those expressing hIDO1 only. In a proteomic analysis, hIDO1-expressing cells exhibited enhanced expression of proteins related to the cell cycle and amino acid metabolism, and decreased expression of proteins related to cell survival. However, cells co-expressing hIDO1 and hIDO2 showed enhanced expression of negative regulators of cell apoptosis compared with those expressing hIDO1 only. Co-expression of hIDO1 and hIDO2 rescued the cell death induced by tryptophan-depletion through hIDO1 activity. Cells expressing only hIDO2 exhibited no marked differences in proteome profiles or cell growth compared with mock-transfectants. Cellular tryptophan metabolic activity and cell death were restored by co-expressing the hIDO2 mutant substituting the histidine 360 residue for alanine. These results demonstrate that hIDO2 plays a novel role as a negative regulator of hIDO1 by competing for heme-binding with hIDO1, and provide information useful for development of therapeutic strategies to control cancer and immunological disorders that target IDO molecules.
Cell Proliferation
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Cell Survival
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Gene Expression
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HEK293 Cells
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Heme/*metabolism
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Humans
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Indoleamine-Pyrrole 2,3,-Dioxygenase/genetics/*metabolism
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Protein Binding
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Tryptophan/*metabolism
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Up-Regulation
2.Molecular cloning and characterization of porcine indoleamine 2, 3-dioxygenase and its expression in various tissues.
Chao CHEN ; Mingfa WEI ; Lu WANG ; Ying XIANG ; Xiangning FU ; Min ZHU
Journal of Huazhong University of Science and Technology (Medical Sciences) 2012;32(4):473-479
In order to confirm the existence of indoleamine 2, 3-dioxygenase (IDO) gene in swine, and to clone the novel gene followed by the molecule structure properties and expression pattern analysis, the porcine mRNA sequences homologous to human IDO were obtained from GenBank database by bioinformatics method. By using RT-PCR, the IDO gene was cloned from porcine endothelial cell line and the accuracy of the nucleic acid sequence was confirmed, and the expression pattern of the gene was detected. The three-dimensional structure model of porcine IDO was built referring to the tertiary structure of human IDO using biological sequence analysis software and database. The results showed that the porcine IDO was identified by sequencing. The nucleotide sequences were confirmed as a novel gene after submitted to Genbank. Porcine IDO was expressed in the lung, thymus, epididymis and anterior chamber with a basic level, however in peripheral blood mononuclear cells (PBMCs) the IDO gene was highly expressed. The three-dimensional structure model of porcine IDO was similar to that of human IDO. It was suggested that identification of the structure information of porcine IDO is essential to further investigate the immunologic function of the gene. Study of IDO on NK cells-mediated xenograft rejection will be a novel therapeutic target for the development of xenotransplantation.
Amino Acid Sequence
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Animals
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Base Sequence
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Cell Line
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Cloning, Molecular
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methods
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Endothelial Cells
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metabolism
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Indoleamine-Pyrrole 2,3,-Dioxygenase
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genetics
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metabolism
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Molecular Sequence Data
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Sequence Alignment
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Swine
3.Effects of Toll-like receptors on indoleamine 2, 3-dioxygenase mRNA levels in human trophoblast HTR-8/SVneo cells.
Wei XU ; Guibo YANG ; Jiazhong DUAN ; Yue WANG ; Wenrong YAO ; Xueqing LIU ; Xuemei CHEN ; Yubin DING ; Yingxiong WANG ; Junlin HE
Journal of Southern Medical University 2013;33(11):1559-1564
OBJECTIVETo study the expression of Toll-like receptors (TLRs) mRNA in human trophoblast HTR-8/SVneo cells and the changes in indoleamine 2,3-dioxygenase (IDO) mRNA expression in response to TLR ligand stimulation.
METHODSThe expressions of TLRs and IDO mRNA in human HTR-8/SVneo cells were tested by RT-PCR, and the changes in IDO mRNA levels after exposure to TLR3, TLR4, TLR7/8, and TLR9 ligands were quantitatively analyzed with real-time PCR.
RESULTSIDO and TLR1-10 mRNAs were expressed in HTR-8/SVneo cells. As the cell culture time extended, IDO mRNA expression level tended to increase within 48 h. After stimulation with the TLR ligands, the expression of TLR-3 mRNA was down-regulated while the expression of TLR-4, 7, 8, and 9 mRNA up-regulated. Stimulation of the cells with poly(I:C) lowered the expression of IDO mRNA while IFN-γ increased its expression.
CONCLUSIONSThe expression of IDO mRNA is associated with the nutrition of the maternal-fetal interface. Stimulation with the TLR ligands affects the expression of IDO and TLR mRNA expressions in the cells, which verifies the functional activity of TLRs and suggests a role of IDO in TLR pathway-dependent antiviral immunity.
Cell Line ; Female ; Humans ; Indoleamine-Pyrrole 2,3,-Dioxygenase ; genetics ; metabolism ; Interferon-gamma ; pharmacology ; Ligands ; Poly I-C ; pharmacology ; RNA, Messenger ; metabolism ; Toll-Like Receptors ; genetics ; metabolism ; Trophoblasts ; cytology ; metabolism
4.Regulatory effect of thymosin α1 on expression of tlr9/ido mRNA in bone marrow mesenchymal stem cells from children with aplastic anemia.
Fang HOU ; Jian-Ming HUANG ; Ge LI
Journal of Experimental Hematology 2010;18(6):1564-1567
The purpose of this study was to explore the regulatory effect of thymosin α1 (Tα1) on expression of TOLL-like receptor 9 (TLR9)/indoleamine2, 3-dioxygenase (ido) mRNA in bone marrow mesenchymal stem cells (MSC) from children with aplastic anemia (AA). Culture system of bone marrow MSC from AA children and normal children in vitro was established, and the effects of Tα1 on expressions of tlr9 mRNA and ido mRNA of MSC from AA children and normal children were determined by RT-PCR. The results showed that the bone marrow MSC from normal children did not express tlr9 and ido mRNA. Bone marrow MSC from children with AA obviously expressed tlr9 mRNA , but did not express ido mRNA; AA children's MSC treated with Tα1 for 18 hours markedly down-regulated tlr9 mRNA expression, but up-regulated ido mRNA expression in the concentration- and time-dependent ways. It is concluded that Tα1 can up-regulate the expression of ido mRNA in bone marrow MSC from children with AA.
Adolescent
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Anemia, Aplastic
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metabolism
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Bone Marrow Cells
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drug effects
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metabolism
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Cells, Cultured
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Child
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Female
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Gene Expression Regulation
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HL-60 Cells
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Humans
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Indoleamine-Pyrrole 2,3,-Dioxygenase
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metabolism
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Male
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Mesenchymal Stromal Cells
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drug effects
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metabolism
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RNA, Messenger
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genetics
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Thymosin
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analogs & derivatives
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pharmacology
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Toll-Like Receptor 9
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metabolism
5.Myeloid differentiation primary response protein 88 blockade upregulates indoleamine 2,3-dioxygenase expression in rheumatoid synovial fibroblasts.
Mi Kyung PARK ; Hye Jwa OH ; Yang Mi HEO ; Eun Mi PARK ; Mi La CHO ; Ho Youn KIM ; Sung Hwan PARK
Experimental & Molecular Medicine 2011;43(8):446-454
Indoleamine 2,3-dioxygenase (IDO) is a key negative regulator of immune responses and has been implicated in tumor tolerance, autoimmune disease and asthma. IDO was detected in the joint synovial tissue in the inflammatory microenvironment of rheumatoid arthritis (RA), but IDO expression in joint synovial tissue is not sufficient to overcome the inflamed synovial environment. This study aimed to unravel the mechanisms involving the failure to activate tolerogenic IDO in the inflamed joint. We demonstrate that both poly (I:C) and lipopolysaccharide (LPS) induce expression of IDO in synovial fibroblasts. However, inflammatory cytokines such as IL-17, TNF-alpha, IL-12, IL-23 and IL-16 did not induce IDO expression. Poly (I:C) appeared to induce higher IDO expression than did LPS. Surprisingly, toll-like receptor (TLR)4-mediated IDO expression was upregulated after depletion of myeloid differentiation primary response protein 88 (MyD88) in synovial fibroblasts using small interfering RNA (siRNA). IDO, TLR3 and TLR4 were highly expressed in synovial tissue of RA patients compared with that of osteoarthritis patients. In addition, RA patients with severe disease activity had higher levels of expression of IDO, TLR3 and TLR4 in the synovium than patients with mild disease activity. These data suggest that upregulation of IDO expression in synovial fibroblasts involves TLR3 and TLR4 activation by microbial constituents. We showed that the mechanisms responsible for IDO regulation primarily involve MyD88 signaling in synovial fibroblasts, as demonstrated by siRNA-mediated knockdown of MyD88.
Adaptor Proteins, Vesicular Transport/genetics/metabolism
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Arthritis, Rheumatoid/*metabolism
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Blotting, Western
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Cells, Cultured
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Fibroblasts/drug effects/*metabolism
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Humans
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Immunohistochemistry
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Indoleamine-Pyrrole 2,3,-Dioxygenase/genetics/*metabolism
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Interleukin-12/pharmacology
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Interleukin-16/pharmacology
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Interleukin-17/pharmacology
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Interleukin-23/pharmacology
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Lipopolysaccharides/pharmacology
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Myeloid Differentiation Factor 88/genetics/*metabolism
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Poly I-C/pharmacology
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Polymerase Chain Reaction
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RNA, Small Interfering/genetics/physiology
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Synovial Membrane/*cytology
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Toll-Like Receptor 4/genetics/metabolism
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Tumor Necrosis Factor-alpha/pharmacology