The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity
碩士 === 國立陽明大學 === 微生物及免疫學研究所 === 99 === In recent years, RIG-I-like receptors (retinoic-acid-inducible gene I-like receptors, RLRs), have been identified that can recognize viral RNA in the cytoplasm and induce the production of type I interferon (IFN). We have established a high-throughput screenin...
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ndltd-TW-099YM0053800152015-10-13T20:37:07Z http://ndltd.ncl.edu.tw/handle/86873877026521339749 The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity 腫瘤原癌基因Pim-1參與調控先天免疫抗病毒機制 Yu-Hsuan Chen 陳宇萱 碩士 國立陽明大學 微生物及免疫學研究所 99 In recent years, RIG-I-like receptors (retinoic-acid-inducible gene I-like receptors, RLRs), have been identified that can recognize viral RNA in the cytoplasm and induce the production of type I interferon (IFN). We have established a high-throughput screening system to discover novel proteins that may be involved in the RLR signaling pathway and selected 15 potential candidates which seemed to affect anti-viral response. We first examined the roles of these candidates in RLR pathway by a reporter assay. The results showed that overexpression of proto-oncogene serine/threonine-protein kinase Pim-1 (Pim-1) enhanced the activation of IFN-β promoter induced by N-RIG (the N-terminal region of RIG-I) in a dose-dependent manner, whereas Pim-1 knockdown reduced the activation. We also examined the effects of Pim-1 on the phosphorylation of endogenous interferon regulatory factor 3 (IRF3) and the expression of IRF3 downstream gene (such as IFN-β, Interferon gamma-induced protein 10 kDa (IP10) and regulated upon activation, normal T-cell expressed, and secreted (RANTES)). All these data conclude that Pim-1 act as a positive regulator for the RIG-I-mediated signaling pathway. Since Pim-1 is a serine/threonine kinase, we therefore generated a kinase dead Pim-1, Pim-1(K67M), and further showed that the mutant Pim-1 significantly lost its regulatory effects, suggesting that Pim-1 regulated RLR signaling through it kinase activity. While Pim-1 strongly enhanced the activation of IFN-β promoter induced by N-RIG, Mitochondrial Antiviral Signaling protein (MAVS) or TANK binding kinase 1 (TBK1), it did not affect the activation of IFN-β promoter induced by Melanoma differentiation-associated gene 5 (MDA5), IRF3-5D (the constitutively active form of IRF3), Inhibitor of nuclear factor kappa-B kinase-beta (IKK-β) or Nuclear factor kappa B (NF-κB(p65)). The results suggest that Pim-1 may up-regulate RLR signaling pathway at a step upstream of MAVS but downstream of RIG-I. Co-immunoprecipitation experiment revealed that Pim-1 could associate with the N-terminal region of RIG-I and extended the half-life of the N-RIG protein. Collectively, these data demonstrate that Pim-1 up-regulates RIG-I-mediated antiviral responses by stabilizing the RIG-I protein through it kinase activity. Lih-Hwa Hwang 黃麗華 2011 學位論文 ; thesis 86 zh-TW |
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碩士 === 國立陽明大學 === 微生物及免疫學研究所 === 99 === In recent years, RIG-I-like receptors (retinoic-acid-inducible gene I-like receptors, RLRs), have been identified that can recognize viral RNA in the cytoplasm and induce the production of type I interferon (IFN). We have established a high-throughput screening system to discover novel proteins that may be involved in the RLR signaling pathway and selected 15 potential candidates which seemed to affect anti-viral response. We first examined the roles of these candidates in RLR pathway by a reporter assay. The results showed that overexpression of proto-oncogene serine/threonine-protein kinase Pim-1 (Pim-1) enhanced the activation of IFN-β promoter induced by N-RIG (the N-terminal region of RIG-I) in a dose-dependent manner, whereas Pim-1 knockdown reduced the activation. We also examined the effects of Pim-1 on the phosphorylation of endogenous interferon regulatory factor 3 (IRF3) and the expression of IRF3 downstream gene (such as IFN-β, Interferon gamma-induced protein 10 kDa (IP10) and regulated upon activation, normal T-cell expressed, and secreted (RANTES)). All these data conclude that Pim-1 act as a positive regulator for the RIG-I-mediated signaling pathway. Since Pim-1 is a serine/threonine kinase, we therefore generated a kinase dead Pim-1, Pim-1(K67M), and further showed that the mutant Pim-1 significantly lost its regulatory effects, suggesting that Pim-1 regulated RLR signaling through it kinase activity. While Pim-1 strongly enhanced the activation of IFN-β promoter induced by N-RIG, Mitochondrial Antiviral Signaling protein (MAVS) or TANK binding kinase 1 (TBK1), it did not affect the activation of IFN-β promoter induced by Melanoma differentiation-associated gene 5 (MDA5), IRF3-5D (the constitutively active form of IRF3), Inhibitor of nuclear factor kappa-B kinase-beta (IKK-β) or Nuclear factor kappa B (NF-κB(p65)). The results suggest that Pim-1 may up-regulate RLR signaling pathway at a step upstream of MAVS but downstream of RIG-I. Co-immunoprecipitation experiment revealed that Pim-1 could associate with the N-terminal region of RIG-I and extended the half-life of the N-RIG protein. Collectively, these data demonstrate that Pim-1 up-regulates RIG-I-mediated antiviral responses by stabilizing the RIG-I protein through it kinase activity.
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author2 |
Lih-Hwa Hwang |
author_facet |
Lih-Hwa Hwang Yu-Hsuan Chen 陳宇萱 |
author |
Yu-Hsuan Chen 陳宇萱 |
spellingShingle |
Yu-Hsuan Chen 陳宇萱 The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity |
author_sort |
Yu-Hsuan Chen |
title |
The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity |
title_short |
The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity |
title_full |
The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity |
title_fullStr |
The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity |
title_full_unstemmed |
The roles of tumor oncogene, Pim-1, involved in RLR-mediated antiviral immunity |
title_sort |
roles of tumor oncogene, pim-1, involved in rlr-mediated antiviral immunity |
publishDate |
2011 |
url |
http://ndltd.ncl.edu.tw/handle/86873877026521339749 |
work_keys_str_mv |
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