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He Q, Sun X, Zhang M, Chu L, Zhao Y, Wu Y, Zhang J, Han X, Guan S, Ding C. Protective effect of baicalin against arsenic trioxide-induced acute hepatic injury in mice through JAK2/STAT3 signaling pathway. Int J Immunopathol Pharmacol 2022; 36:20587384211073397. [PMID: 35088608 PMCID: PMC8801635 DOI: 10.1177/20587384211073397] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Baicalin (BA) is a kind of flavonoid that is isolated from Scutellaria baicalensis Georgi, which has been verified to have hepatoprotective effects in some diseases. However, the role of BA in acute hepatic injury induced by arsenic trioxide (ATO) remains unclear. The aim of this study was to investigate the protective action of BA on acute hepatic injury induced by ATO and to probe its possible mechanism. Mice were pretreated with BA (50, 100 mg/kg) by gavage. After 7 h, ATO (7.5 mg/kg) was injected intraperitoneally to induce liver injury. After 7 days of treatment, serum and hepatic specimens were collected and assayed to evaluate the hepatoprotective effect of BA. Pathological sections and the liver function index indicated that ATO caused significant liver injury. The fluorescence of reactive oxygen species and oxidative stress indicators showed that ATO also increased oxidative stress. The inflammatory markers in ATO-induced mice also increased significantly. Staining of the terminal deoxynucleotidyl transferase dUTP nick end labeling and apoptotic factor assay showed that apoptosis increased. However, with BA pretreatment, these changes were significantly weakened. In addition, BA treatment promoted the expression of proteins related to the JAK2/STAT3 signaling pathway. The results suggest that BA can ameliorate acute ATO-induced hepatic injury in mice, which is related to the inhibition of oxidative stress, thereby reducing inflammation and apoptosis. The mechanism of this protection is potentially related to the JAK2/STAT3 signaling pathway.
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Affiliation(s)
- Qianqian He
- School of Pharmacy, Hebei University of Chinese Medicine, Shijiazhuang, China
| | - Xiaoqi Sun
- School of Pharmacy, Hebei University of Chinese Medicine, Shijiazhuang, China
| | - Muqing Zhang
- Affiliated Hospital, Hebei University of Chinese Medicine, Shijiazhuang, China
- College of Integrative Medicine, Hebei University of Chinese Medicine, Shijiazhuang, China
| | - Li Chu
- School of Pharmacy, Hebei University of Chinese Medicine, Shijiazhuang, China
- Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns, Shijiazhuang, China
| | - Yang Zhao
- The Fourth Hospital of Hebei Medical University, Shijiazhuang, China
| | - Yongchao Wu
- The Fourth Hospital of Hebei Medical University, Shijiazhuang, China
| | - Jianping Zhang
- Hebei Key Laboratory of Integrative Medicine on Liver-Kidney Patterns, Shijiazhuang, China
- School of Basic Medicine, Hebei University of Chinese Medicine, Shijiazhuang, China
| | - Xue Han
- School of Pharmacy, Hebei University of Chinese Medicine, Shijiazhuang, China
- Affiliated Hospital, Hebei University of Chinese Medicine, Shijiazhuang, China
| | - Shengjiang Guan
- Affiliated Hospital, Hebei University of Chinese Medicine, Shijiazhuang, China
- School of Basic Medicine, Hebei University of Chinese Medicine, Shijiazhuang, China
- Shengjiang Guan, Affiliated Hospital, Hebei University of Chinese Medicine, No. 3, Xingyuan Road, Luquan Economic Development Zone, Luquan District, Shijiazhuang, Hebei 050011, China.
| | - Chao Ding
- Department of Cardiology, Bethune International Peace Hospital of PLA, Shijiazhuang, China
- Chao Ding, Department of Cardiology, Bethune International Peace Hospital of PLA, Shijiazhuang, Hebei 050011, China. Email
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Strous GJ, Almeida ADS, Putters J, Schantl J, Sedek M, Slotman JA, Nespital T, Hassink GC, Mol JA. Growth Hormone Receptor Regulation in Cancer and Chronic Diseases. Front Endocrinol (Lausanne) 2020; 11:597573. [PMID: 33312162 PMCID: PMC7708378 DOI: 10.3389/fendo.2020.597573] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/21/2020] [Accepted: 10/14/2020] [Indexed: 12/14/2022] Open
Abstract
The GHR signaling pathway plays important roles in growth, metabolism, cell cycle control, immunity, homeostatic processes, and chemoresistance via both the JAK/STAT and the SRC pathways. Dysregulation of GHR signaling is associated with various diseases and chronic conditions such as acromegaly, cancer, aging, metabolic disease, fibroses, inflammation and autoimmunity. Numerous studies entailing the GHR signaling pathway have been conducted for various cancers. Diverse factors mediate the up- or down-regulation of GHR signaling through post-translational modifications. Of the numerous modifications, ubiquitination and deubiquitination are prominent events. Ubiquitination by E3 ligase attaches ubiquitins to target proteins and induces proteasomal degradation or starts the sequence of events that leads to endocytosis and lysosomal degradation. In this review, we discuss the role of first line effectors that act directly on the GHR at the cell surface including ADAM17, JAK2, SRC family member Lyn, Ubc13/CHIP, proteasome, βTrCP, CK2, STAT5b, and SOCS2. Activity of all, except JAK2, Lyn and STAT5b, counteract GHR signaling. Loss of their function increases the GH-induced signaling in favor of aging and certain chronic diseases, exemplified by increased lung cancer risk in case of a mutation in the SOCS2-GHR interaction site. Insight in their roles in GHR signaling can be applied for cancer and other therapeutic strategies.
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Affiliation(s)
- Ger J. Strous
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
- BIMINI Biotech B.V., Leiden, Netherlands
| | - Ana Da Silva Almeida
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Joyce Putters
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Julia Schantl
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Magdalena Sedek
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Johan A. Slotman
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Tobias Nespital
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Gerco C. Hassink
- Department of Cell Biology, Centre for Molecular Medicine, University Medical Centre Utrecht, Utrecht, Netherlands
| | - Jan A. Mol
- Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Utrecht, Netherlands
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Tolomeo M, Meli M, Grimaudo S. STAT5 and STAT5 Inhibitors in Hematological Malignancies. Anticancer Agents Med Chem 2019; 19:2036-2046. [PMID: 31490767 DOI: 10.2174/1871520619666190906160848] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2019] [Revised: 07/09/2019] [Accepted: 07/18/2019] [Indexed: 11/22/2022]
Abstract
The JAK-STAT pathway is an important physiologic regulator of different cellular functions including proliferation, apoptosis, differentiation, and immunological responses. Out of six different STAT proteins, STAT5 plays its main role in hematopoiesis and constitutive STAT5 activation seems to be a key event in the pathogenesis of several hematological malignancies. This has led many researchers to develop compounds capable of inhibiting STAT5 activation or interfering with its functions. Several anti-STAT5 molecules have shown potent STAT5 inhibitory activity in vitro. However, compared to the large amount of clinical studies with JAK inhibitors that are currently widely used in the clinics to treat myeloproliferative disorders, the clinical trials with STAT5 inhibitors are very limited. At present, a few STAT5 inhibitors are in phase I or II clinical trials for the treatment of leukemias and graft vs host disease. These studies seem to indicate that such compounds could be well tolerated and useful in reducing the occurrence of resistance to tyrosine kinase inhibitors in chronic myeloid leukemia. Of interest, STAT5 seems to play an important role in the regulation of hematopoietic stem cell self-renewal suggesting that combination therapies including STAT5 inhibitors can erode the cancer stem cell pool and possibly open the way for the complete cancer eradication. In this review, we discuss the implication of STAT5 in hematological malignancies and the results obtained with the novel STAT5 inhibitors.
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Affiliation(s)
- Manlio Tolomeo
- Department of Health Promotion Sciences, Maternal and Infant Care, Internal Medicine and Medical Specialties, University of Palermo, Palermo, Italy
| | - Maria Meli
- Department of Health Promotion Sciences, Maternal and Infant Care, Internal Medicine and Medical Specialties, University of Palermo, Palermo, Italy
| | - Stefania Grimaudo
- Department of Health Promotion Sciences, Maternal and Infant Care, Internal Medicine and Medical Specialties, University of Palermo, Palermo, Italy
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Chen Y, Zhu Y, Sheng Y, Xiao J, Xiao Y, Cheng N, Chai Y, Wu X, Zhang S, Xiang T. SIRT1 downregulated FGB expression to inhibit RCC tumorigenesis by destabilizing STAT3. Exp Cell Res 2019; 382:111466. [PMID: 31201813 DOI: 10.1016/j.yexcr.2019.06.011] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2019] [Revised: 06/10/2019] [Accepted: 06/11/2019] [Indexed: 12/21/2022]
Abstract
Renal cell carcinoma (RCC) is one of the common lethal urologic tumors. Recent studies revealed that SIRT1 might function as a tumor suppressor during the progression of RCC. In addition, studies showed that FGB expression was abnormally upregulated in RCC and related to the progress of RCC. This study aimed to define the function of SIRT1 and underlying mechanism in the RCC progression. The expression of SIRT1 and FGB in RCC specimens and cells were detected by immunoblotting and immunostaining. Luciferase reporter assay was performed to confirm FGB as the target gene of STAT3. Other methods including stable transfection, co-immunoprecipitation, Western blot, and in vitro and in vivo proliferation assays were also performed. Our results showed that SIRT1 expression was downregulated in RCC tissues compared to adjacent normal tissues and relatively high expression of SIRT1 conferred a better prognosis for patients. Next, we showed that SIRT1 overexpression inhibited RCC tumorigenesis both in vitro and in vivo. In addition, FGB expression was upregulated in RCC tissues and overexpressing SIRT1 reduced FGB expression levels. Furthermore, inhibition of RCC proliferation by SIRT1 overexpression was rescued by FGB overexpression, indicating that SIRT1 inhibited RCC proliferation by repressing FGB expression. Mechanistically, we confirmed that FGB was the target gene of STAT3, and SIRT1 repressed the expression of FGB by deacetylation of STAT3, leading to STAT3 destabilization and degradation. SIRT1 inhibited RCC tumorigenesis by downregulating FGB expression, and this novel SIRT1-STAT3-FGB axis provided a potential target for RCC therapy.
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Affiliation(s)
- Yanbing Chen
- Department of Nephrology, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China
| | - Ying Zhu
- Department of Infectious Disease, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China
| | - Yanling Sheng
- Department of Ultrasound, The Affiliated Hospital of Jiangxi University of Traditional Chinese Medicine, Nanchang, 330006, Jiangxi Province, China
| | - Juhua Xiao
- Department of Ultrasound, Jiangxi Provincial Maternal and Child Health Hospital, Nanchang, Jiangxi Province, Jiangxi, 330006, China
| | - Yu Xiao
- Department of General Surgery, Jiangxi Provincial Children's Hospital, Nanchang, Jiangxi Province, Jiangxi, 330006, China
| | - Na Cheng
- Department of Infectious Disease, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China
| | - Yong Chai
- Department of General Surgery, Jiangxi Provincial Children's Hospital, Nanchang, Jiangxi Province, Jiangxi, 330006, China
| | - Xiaoping Wu
- Department of Infectious Disease, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China
| | - Shouhua Zhang
- Department of General Surgery, Jiangxi Provincial Children's Hospital, Nanchang, Jiangxi Province, Jiangxi, 330006, China.
| | - Tianxin Xiang
- Department of Infectious Disease, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, 330006, China.
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Circular RNA 0039411 Is Involved in Neodymium Oxide-induced Inflammation and Antiproliferation in a Human Bronchial Epithelial Cell Line via Sponging miR-93-5p. Toxicol Sci 2019; 170:69-81. [DOI: 10.1093/toxsci/kfz074] [Citation(s) in RCA: 30] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/22/2023] Open
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Nabavi SM, Ahmed T, Nawaz M, Devi KP, Balan DJ, Pittalà V, Argüelles-Castilla S, Testai L, Khan H, Sureda A, de Oliveira MR, Vacca RA, Xu S, Yousefi B, Curti V, Daglia M, Sobarzo-Sánchez E, Filosa R, Nabavi SF, Majidinia M, Dehpour AR, Shirooie S. Targeting STATs in neuroinflammation: The road less traveled! Pharmacol Res 2018; 141:73-84. [PMID: 30550953 DOI: 10.1016/j.phrs.2018.12.004] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/23/2018] [Revised: 12/01/2018] [Accepted: 12/10/2018] [Indexed: 12/16/2022]
Abstract
JAK/STAT transduction pathway is a highly conserved pathway implicated in regulating cellular proliferation, differentiation, survival and apoptosis. Dysregulation of this pathway is involved in the onset of autoimmune, haematological, oncological, metabolic and neurological diseases. Over the last few years, the research of anti-neuroinflammatory agents has gained considerable attention. The ability to diminish the STAT-induced transcription of inflammatory genes is documented for both natural compounds (such as polyphenols) and chemical drugs. Among polyphenols, quercetin and curcumin directly inhibit STAT, while Berberis vulgaris L. and Sophora alopecuroides L extracts act indirectly. Also, the Food and Drug Administration has approved several JAK/STAT inhibitors (direct or indirect) for treating inflammatory diseases, indicating STAT can be considered as a therapeutic target for neuroinflammatory pathologies. Considering the encouraging data obtained so far, clinical trials are warranted to demonstrate the effectiveness and potential use in the clinical practice of STAT inhibitors to treat inflammation-associated neurodegenerative pathologies.
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Affiliation(s)
- Seyed Mohammad Nabavi
- Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
| | - Touqeer Ahmed
- Neurobiology Laboratory, Department of Healthcare Biotechnology, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Sector H-12, Islamabad, 44000, Pakistan
| | - Maheen Nawaz
- Neurobiology Laboratory, Department of Healthcare Biotechnology, Atta-ur-Rahman School of Applied Biosciences, National University of Sciences and Technology, Sector H-12, Islamabad, 44000, Pakistan
| | - Kasi Pandima Devi
- Department of Biotechnology, Alagappa University (Science Campus), Karaikudi 630 003, Tamil Nadu, India
| | - Devasahayam Jaya Balan
- Department of Biotechnology, Alagappa University (Science Campus), Karaikudi 630 003, Tamil Nadu, India
| | - Valeria Pittalà
- Department of Drug Sciences, University of Catania, Viale A. Doria 6, 95125, Catania, Italy
| | | | - Lara Testai
- Department of Pharmacy, University of Pisa, Pisa, via Bonanno 6 - 56126, Pisa, Italy
| | - Haroon Khan
- Department of Pharmacy, Abdul Wali Khan University, Mardan 23200, Pakistan
| | - Antoni Sureda
- Research Group on Community Nutrition and Oxidative Stress and CIBEROBN (Physiopathology of Obesity and Nutrition), University of Balearic Islands, E-07122 Palma de Mallorca, Spain.
| | - Marcos Roberto de Oliveira
- Department of Chemistry/ICET, Federal University of Mato Grosso (UFMT), Av. Fernando Corrêa da Costa, 2367, Cuiaba, MT, 78060-900, Brazil
| | - Rosa Anna Vacca
- Institute of Biomembranes, Bioenergetics and Molecular Biotechnologies, National Council of Research, I-70126, Bari, Italy
| | - Suowen Xu
- University of Rochester, Aab Cardiovascular Research Institute, Rochester, NY, 14623, USA
| | - Bahman Yousefi
- Molecular Medicine Research Center, Tabriz University of Medical Sciences, Tabriz, Iran; Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Valeria Curti
- Department of Drug Sciences, University of Pavia, Pavia, Italy
| | - Maria Daglia
- Department of Drug Sciences, University of Pavia, Pavia, Italy
| | - Eduardo Sobarzo-Sánchez
- Laboratory of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Santiago de Compostela, 15782, Spain; Instituto de Investigación e Innovación en Salud, Facultad de Ciencias de la Salud, Universidad Central de Chile, Santiago, Chile
| | - Rosanna Filosa
- Consorzio Sannio Tech, Appia Str, Apollosa, BN 82030, Italy
| | - Seyed Fazel Nabavi
- Pharmaceutical Sciences Research Center, Kermanshah University of Medical Sciences, Kermanshah, Iran; Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
| | - Maryam Majidinia
- Solid Tumor Research Center, Urmia University of Medical Sciences, Urmia, Iran
| | - Ahmad Reza Dehpour
- Department of Pharmacology, Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Iran; Experimental Medicine Research Center, Tehran University of Medical Sciences, Tehran, Iran
| | - Samira Shirooie
- Department of Pharmacology, Faculty of Pharmacy, Kermanshah University of Medical Sciences, Kermanshah, Iran
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Chaudhary O, Narayan V, Lelis F, Linz B, Watkins M, Veazey R, Aldovini A. Inhibition of p38 MAPK in combination with ART reduces SIV-induced immune activation and provides additional protection from immune system deterioration. PLoS Pathog 2018; 14:e1007268. [PMID: 30161247 PMCID: PMC6135519 DOI: 10.1371/journal.ppat.1007268] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2018] [Revised: 09/12/2018] [Accepted: 08/08/2018] [Indexed: 12/12/2022] Open
Abstract
Differences in immune activation were identified as the most significant difference between AIDS-susceptible and resistant species. p38 MAPK, activated in HIV infection, is key to induction of interferon-stimulated genes and cytokine-mediated inflammation and is associated with some of the pathology produced by HIV or SIV infection in AIDS-susceptible primates. As small molecule p38 MAPK inhibitors are being tested in human trials for inflammatory diseases, we evaluated the effects of treating SIV-infected macaques with the p38 MAPK inhibitor PH-797804 in conjunction with ART. PH-797804 had no side effects, did not impact negatively the antiviral immune response and, used alone, had no significant effect on levels of immune activation and did not reduced the viremia. When administered with ART, it significantly reduced numerous immune activation markers compared to ART alone. CD38+/HLA-DR+ and Ki-67+ T-cell percentages in blood, lymph node and rectal CD4+ and CD8+ T cells, PD-1 expression in CD8+ T cells and plasma levels of IFNα, IFNγ, TNFα, IL-6, IP-10, sCD163 and C-reactive protein were all significantly reduced. Significant preservation of CD4+, CD4+ central memory, CD4+/IL-22+ and CD4+/IL-17+ T-cell percentages and improvement of Th17/Treg ratio in blood and rectal mucosa were also observed. Importantly, the addition of PH-797804 to ART initiated during chronic SIV infection reduced immune activation and restored immune system parameters to the levels observed when ART was initiated on week 1 after infection. After ART interruption, viremia rebounded in a similar fashion in all groups, regardless of when ART was initiated. We concluded that the inhibitor PH-797804 significantly reduced, even if did not normalized, the immune activation parameters evaluated during ART treatment, improved preservation of critical populations of the immune system targeted by SIV, and increased the efficacy of ART treatment initiated in chronic infection to levels similar to those observed when initiated in acute infection but did not affect positively or negatively viral reservoirs. The hallmark of Human Immunodeficiency Virus and Simian Immunodeficiency Virus infection in disease-susceptible species is the progressive decline of the CD4+ T cell population and heightened immune activation, which by itself can contribute to CD4+ T-cell death. The cellular pathway regulated by p38 MAPK, which is activated in HIV and SIV infection, can contribute significantly to immune activation. We tested in SIV-infected macaques a p38 MAPK inhibitor in combination with anti-retroviral therapy. This drug is already being evaluated in humans for treatment of immune activation associated with other diseases. We found that, when combined with antiretroviral therapy, the inhibitor PH-797804 significantly reduced a few parameters of SIV-induced immune activation and improved preservation of critical populations of the immune system targeted by SIV, but did not modulate viral reservoirs. Importantly, the addition of the inhibitor to anti-retroviral therapy during the chronic phase of the infection, which is the time when most HIV-infected individuals initiate treatment, permitted a more significant preservation of the immune system compared to antiretroviral therapy alone that was similar to that observed when anti-retroviral therapy was initiated in the acute phase of the infection, which rarely occurs in HIV infection.
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Affiliation(s)
- Omkar Chaudhary
- Boston Children’s Hospital, Department of Medicine, and Harvard Medical School, Department of Pediatrics, Boston MA, United States of America
| | - Vivek Narayan
- Boston Children’s Hospital, Department of Medicine, and Harvard Medical School, Department of Pediatrics, Boston MA, United States of America
| | - Felipe Lelis
- Boston Children’s Hospital, Department of Medicine, and Harvard Medical School, Department of Pediatrics, Boston MA, United States of America
| | - Brandon Linz
- Boston Children’s Hospital, Department of Medicine, and Harvard Medical School, Department of Pediatrics, Boston MA, United States of America
| | - Meagan Watkins
- Tulane National Primate Research Center, Division of Comparative Pathology, Covington LA, United States of America
| | - Ronald Veazey
- Tulane National Primate Research Center, Division of Comparative Pathology, Covington LA, United States of America
| | - Anna Aldovini
- Boston Children’s Hospital, Department of Medicine, and Harvard Medical School, Department of Pediatrics, Boston MA, United States of America
- * E-mail:
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Dynamic Changes in the Splenic Transcriptome of Chickens during the Early Infection and Progress of Marek's Disease. Sci Rep 2017; 7:11648. [PMID: 28912500 PMCID: PMC5599560 DOI: 10.1038/s41598-017-11304-y] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2016] [Accepted: 08/22/2017] [Indexed: 01/18/2023] Open
Abstract
Gallid alphaherpesvirus 2 (GaHV2) is an oncogenic avian herpesvirus inducing Marek’s disease (MD) and rapid-onset T-cell lymphomas. To reveal molecular events in MD pathogenesis and tumorigenesis, the dynamic splenic transcriptome of GaHV2-infected chickens during early infection and pathogenic phases has been determined utilizing RNA-seq. Based on the significant differentially expressed genes (DEGs), analysis of gene ontology, KEGG pathway and protein-protein interaction network has demonstrated that the molecular events happening during GaHV2 infection are highly relevant to the disease course. In the ‘Cornell Model’ description of MD, innate immune responses and inflammatory responses were established at early cytolytic phase but persisted until lymphoma formation. Humoral immunity in contrast began to play a role firstly in the intestinal system and started at late cytolytic phase. Neurological damage caused by GaHV2 is first seen in early cytolytic phase and is then sustained throughout the following phases over a long time period. During the proliferative phase many pathways associated with transcription and/or translation were significantly enriched, reflecting the cell transformation and lymphoma formation. Our work provides an overall view of host responses to GaHV2 infection and offers a meaningful basis for further studies of MD biology.
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Dai YJ, Hui KM, Zhang YH, Liu Y, Wang YQ, Zhao LJ, Lin L, Chai LQ, Wei S, Lan JF. Three STATs are involved in the regulation of the expression of antimicrobial peptides in the triangle sail mussel, Hyriopsis cumingii. FISH & SHELLFISH IMMUNOLOGY 2017; 63:181-188. [PMID: 28214598 DOI: 10.1016/j.fsi.2017.02.012] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/07/2016] [Revised: 02/06/2017] [Accepted: 02/10/2017] [Indexed: 05/25/2023]
Abstract
Janus kinase (Jak) and signal transducers and activators of transcription (STAT) signaling pathway is associated in antiviral and antibacterial immune response. Previous studies primarily investigated the function of STATs in mammals. For most invertebrates, only one STAT was found in each species, such as STAT92E was found in Drosophila melanogaster. The studies, which focus on the functional difference between various STATs in the same species of invertebrate, are limited. In the present study, three STATs (HcSTAT1, HcSTAT2 and HcSTAT3) were identified in triangle shell pearl mussel, Hyriopsis cumingii. Phylogenetic analysis showed that HcSTAT1 and HcSTAT3 were clustered with Homo sapiens STAT5, and HcSTAT2 was clustered with Pinctada fucata STAT and Crassostea gigas STAT6. All three STATs could be detected in all tested tissues (hemocytes, hepatopancreas, gill, mantle and foot), and were induced expression when challenged with Staphylococcus aureus or Aeromonas hydrophilia in hemocytes and hepatopancreas. HcSTAT1 regulated the expression of HcDef, HcWAP, HcThe and HcTNF. The expression of HcWAP and HcTNF was down-regulated in HcSTAT2-RNAi mussel. And HcSTAT3 affected the expression of HcTNF. The study is the first report of different functions in antibacterial immune responses between STATs in mollusks.
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Affiliation(s)
- Yun-Jia Dai
- State Key Laboratory of Cotton Biology, School of Life Sciences Henan University, Kaifeng 475004, China; Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China
| | - Kai-Min Hui
- Jiangsu Key Laboratory for Biodiversity & Biotechnology and Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Life Sciences, Nanjing Normal University, Nanjing 210046, China
| | - Ying-Hao Zhang
- Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China
| | - Yan Liu
- Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China
| | - Yu-Qing Wang
- Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China
| | - Li-Juan Zhao
- Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China; Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding, College of Animal Sciences and Technology, Zhongkai University of Agriculture and Engineering, Guangzhou, Guangdong 510225, China
| | - Li Lin
- Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China; Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding, College of Animal Sciences and Technology, Zhongkai University of Agriculture and Engineering, Guangzhou, Guangdong 510225, China
| | - Lian-Qin Chai
- State Key Laboratory of Cotton Biology, School of Life Sciences Henan University, Kaifeng 475004, China.
| | - Shun Wei
- Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China.
| | - Jiang-Feng Lan
- State Key Laboratory of Cotton Biology, School of Life Sciences Henan University, Kaifeng 475004, China; Department of Aquatic Animal Medicine, Research Center for Marine Biology, College of Fisheries, Huazhong Agricultural University, Wuhan 430070, China; Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangzhou 510380, China.
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Beutel O, Roder F, Birkholz O, Rickert C, Steinhoff HJ, Grzybek M, Coskun Ü, Piehler J. Two-Dimensional Trap for Ultrasensitive Quantification of Transient Protein Interactions. ACS NANO 2015; 9:9783-9791. [PMID: 26331529 DOI: 10.1021/acsnano.5b02696] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/05/2023]
Abstract
We present an ultrasensitive technique for quantitative protein-protein interaction analysis in a two-dimensional format based on phase-separated, micropatterned membranes. Interactions between proteins captured to lipid probes via an affinity tag trigger partitioning into the liquid-ordered phase, which is readily quantified by fluorescence imaging. Based on a calibration with well-defined low-affinity protein-protein interactions, equilibrium dissociation constants >1 mM were quantified. Direct capturing of proteins from mammalian cell lysates enabled us to detect homo- and heterodimerization of signal transducer and activator of transcription proteins. Using the epidermal growth factor receptor (EGFR) as a model system, quantification of low-affinity interactions between different receptor domains contributing to EGFR dimerization was achieved. By exploitation of specific features of the membrane-based assay, the regulation of EGFR dimerization by lipids was demonstrated.
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Affiliation(s)
- Oliver Beutel
- Department of Biology, University of Osnabrück , 49074 Osnabrück, Germany
| | - Friedrich Roder
- Department of Biology, University of Osnabrück , 49074 Osnabrück, Germany
| | - Oliver Birkholz
- Department of Biology, University of Osnabrück , 49074 Osnabrück, Germany
| | - Christian Rickert
- Department of Physics, University of Osnabrück , 49076 Osnabrück, Germany
| | | | - Michał Grzybek
- Paul Langerhans Institute Dresden of the Helmholtz Centre Munich at the University Clinic Carl Gustav Carus TU Dresden , 01307 Dresden, Germany
- German Center for Diabetes Research (DZD) , 85764 Neuherberg, Germany
| | - Ünal Coskun
- Paul Langerhans Institute Dresden of the Helmholtz Centre Munich at the University Clinic Carl Gustav Carus TU Dresden , 01307 Dresden, Germany
- German Center for Diabetes Research (DZD) , 85764 Neuherberg, Germany
| | - Jacob Piehler
- Department of Biology, University of Osnabrück , 49074 Osnabrück, Germany
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Abroun S, Saki N, Ahmadvand M, Asghari F, Salari F, Rahim F. STATs: An Old Story, Yet Mesmerizing. CELL JOURNAL 2015; 17:395-411. [PMID: 26464811 PMCID: PMC4601860 DOI: 10.22074/cellj.2015.1] [Citation(s) in RCA: 57] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/02/2013] [Accepted: 08/07/2014] [Indexed: 01/01/2023]
Abstract
Signal transducers and activators of transcription (STATs) are cytoplasmic transcription factors that have a key role in cell fate. STATs, a protein family comprised of
seven members, are proteins which are latent cytoplasmic transcription factors that
convey signals from the cell surface to the nucleus through activation by cytokines
and growth factors. The signaling pathways have diverse biological functions that
include roles in cell differentiation, proliferation, development, apoptosis, and inflammation which place them at the center of a very active area of research. In this review we explain Janus kinase (JAK)/STAT signaling and focus on STAT3, which is
transient from cytoplasm to nucleus after phosphorylation. This procedure controls
fundamental biological processes by regulating nuclear genes controlling cell proliferation, survival, and development. In some hematopoietic disorders and cancers,
overexpression and activation of STAT3 result in high proliferation, suppression of
cell differentiation and inhibition of cell maturation. This article focuses on STAT3
and its role in malignancy, in addition to the role of microRNAs (miRNAs) on STAT3
activation in certain cancers.
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Affiliation(s)
- Saeid Abroun
- Department of Hematology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
| | - Najmaldin Saki
- Health Research Institute, Research Center of Thalassemia and Hemoglobinopathy, Jundishapur University of Medical Sciences, Ahvaz, Iran
| | - Mohammad Ahmadvand
- Department of Hematology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
| | - Farahnaz Asghari
- Department of Medicine II, Division of Gastroenterology, University of Rostock, E.Heydemann-Strasse 6, Rostock, Germany
| | - Fatemeh Salari
- Health Research Institute, Research Center of Thalassemia and Hemoglobinopathy, Jundishapur University of Medical Sciences, Ahvaz, Iran
| | - Fakher Rahim
- Health Research Institute, Hearing Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran
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Moravcová S, Červená K, Pačesová D, Bendová Z. Identification of STAT3 and STAT5 proteins in the rat suprachiasmatic nucleus and the Day/Night difference in astrocytic STAT3 phosphorylation in response to lipopolysaccharide. J Neurosci Res 2015; 94:99-108. [PMID: 26420542 DOI: 10.1002/jnr.23673] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2015] [Revised: 09/01/2015] [Accepted: 09/08/2015] [Indexed: 12/12/2022]
Abstract
Signal transducers and activators of transcription (STAT) proteins regulate many aspects of cellular physiology from growth and differentiations to immune responses. Using immunohistochemistry, we show the daily rhythm of STAT3 protein in the rat suprachiasmatic nucleus (SCN), with low but significant amplitude peaking in the morning. We also reveal the strong expression of STAT5A in astrocytes of the SCN and the STAT5B signal in nonastrocytic cells. Administration of lipopolysaccharide (LPS) acutely induced phosphorylation of STAT3 on Tyr705 during both the day and the night and induced phosphorylation on Ser727 but only after the daytime application. The LPS-induced phospho-STAT3 (Tyr705) remained elevated for 24 hr after the daytime application but declined within 8 hr when LPS was applied at night.
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Affiliation(s)
- Simona Moravcová
- Department of Physiology, Faculty of Science, Charles University in Prague, Prague, Czech Republic
| | - Kateřina Červená
- Department of Physiology, Faculty of Science, Charles University in Prague, Prague, Czech Republic
| | - Dominika Pačesová
- Department of Physiology, Faculty of Science, Charles University in Prague, Prague, Czech Republic
| | - Zdeňka Bendová
- Department of Physiology, Faculty of Science, Charles University in Prague, Prague, Czech Republic
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13
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Zhou Y, Tian L, Zhang YC, Guo BF, Zhou QW. Apoptotic effects of psiRNA-STAT3 on 4T1 breast cancer cells in vitro. Asian Pac J Cancer Prev 2015; 15:6977-82. [PMID: 25169471 DOI: 10.7314/apjcp.2014.15.16.6977] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022] Open
Abstract
BACKGROUND The aim of this study was to investigate the effect of a Lipofectamine2000 (Life2000) Transfection Reagent transfected psiRNA-STAT3 plasmid on 4T1 breast cancer cells. MATERIALS AND METHODS MTT was used to detect the cell proliferation of breast cancer 4T1 cells at different periods (0h, 6h, 8h, 10h); the cell cycle was assessed by flow cytometry; variation of apoptosis and mitochondrial membrane potential was observed under a fluorescence microscope; immunohistochemical staining was used to determine the expression of caspase-3 and cyclin-D1 protein. RESULTS An obvious effect of inhibition to 4T1 cancer cells could be observed at 8h after the psiRNA-STAT3 was transfected. Typical alterations of apoptotic morphological features were visible in the psiRNA-STAT3 treatment group. Mitochondrial membrane potential decreased significantly, the number of cells was increased in G0/G1 phase, and the number of cells was decreased in S phase, and the data were statistically significant (p<0.05), compared with the Scramble and Mock groups. Expression of caspase-3 protein was increased significantly, while that of cyclin D1 was significantly decreased. CONCLUSIONS Life2000 transfected psiRNA-STAT3 plasmid can inhibit 4T1 tumor cell proliferation and promote apoptosis of 4T1 tumor cells, which process depends on the regulation of expression of cyclin D1 and caspase-3 protein.
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Affiliation(s)
- Yue Zhou
- School of Pharmacy, 2Department of Breast Surgery , The Second Clinical Hospital, 3Department of Plastic Surgery, the China- Japan Union Hospital, 4Department of Biology and Medical Engineering, Institute of Regenerative Medicine, Jilin University, Changchun, China E-mail : ,
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14
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Wei W, Lewis MT. Identifying and targeting tumor-initiating cells in the treatment of breast cancer. Endocr Relat Cancer 2015; 22:R135-55. [PMID: 25876646 PMCID: PMC4447610 DOI: 10.1530/erc-14-0447] [Citation(s) in RCA: 40] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 04/07/2015] [Indexed: 01/05/2023]
Abstract
Breast cancer is the most common cancer in women (excluding skin cancer), and it is the second leading cause of cancer-related deaths. Although conventional and targeted therapies have improved survival rates, there are still considerable challenges in treating breast cancer, including treatment resistance, disease recurrence, and metastasis. Treatment resistance can be either de novo - because of traits that tumor cells possess before treatment - or acquired - because of traits that tumor cells gain in response to treatment. A recently proposed mechanism of de novo resistance invokes the existence of a specialized subset of cancer cells defined as tumor-initiating cells (TICs), or cancer stem cells (CSCs). TICs have the capacity to self-renew and to generate new tumors that consist entirely of clonally derived cell types present in the parental tumor. There are data to suggest that TICs are resistant to many conventional cancer therapies and that they can survive treatment in spite of dramatic shrinkage of the tumor. Residual TICs can then eventually regrow, which results in disease relapse. It has also been hypothesized that TIC may be responsible for metastatic disease. If these hypotheses are correct, targeting TICs may be imperative for achieving a cure. In the present review, we discuss evidence for breast TICs and their apparent resistance to conventional chemotherapy and radiotherapy as well as to various targeted therapies. We also address the potential impact of breast TIC plasticity and metastatic potential on therapeutic strategies. Finally, we describe several genes and signaling pathways that appear to be important for TIC function and may represent promising therapeutic targets.
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Affiliation(s)
- Wei Wei
- Baylor College of MedicineLester and Sue Smith Breast Center, Houston, Texas, USADepartments of Molecular and Cellular BiologyRadiologyBaylor College of Medicine, One Baylor Plaza, BCM600, Room N1210, Houston, Texas 77030, USA Baylor College of MedicineLester and Sue Smith Breast Center, Houston, Texas, USADepartments of Molecular and Cellular BiologyRadiologyBaylor College of Medicine, One Baylor Plaza, BCM600, Room N1210, Houston, Texas 77030, USA
| | - Michael T Lewis
- Baylor College of MedicineLester and Sue Smith Breast Center, Houston, Texas, USADepartments of Molecular and Cellular BiologyRadiologyBaylor College of Medicine, One Baylor Plaza, BCM600, Room N1210, Houston, Texas 77030, USA Baylor College of MedicineLester and Sue Smith Breast Center, Houston, Texas, USADepartments of Molecular and Cellular BiologyRadiologyBaylor College of Medicine, One Baylor Plaza, BCM600, Room N1210, Houston, Texas 77030, USA Baylor College of MedicineLester and Sue Smith Breast Center, Houston, Texas, USADepartments of Molecular and Cellular BiologyRadiologyBaylor College of Medicine, One Baylor Plaza, BCM600, Room N1210, Houston, Texas 77030, USA
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15
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Wei W, Tweardy DJ, Zhang M, Zhang X, Landua J, Petrovic I, Bu W, Roarty K, Hilsenbeck SG, Rosen JM, Lewis MT. STAT3 signaling is activated preferentially in tumor-initiating cells in claudin-low models of human breast cancer. Stem Cells 2015; 32:2571-82. [PMID: 24891218 DOI: 10.1002/stem.1752] [Citation(s) in RCA: 83] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2013] [Revised: 04/16/2014] [Accepted: 05/03/2014] [Indexed: 12/31/2022]
Abstract
In breast cancer, a subset of tumor-initiating cells (TIC) or "cancer stem cells" are thought to be responsible for tumor maintenance, treatment resistance, and disease recurrence. While current breast cancer stem cell markers (e.g., CD44(high) /CD24(low/neg) , ALDH positive) have allowed enrichment for such cells, they are not universally expressed and may actually identify distinct TIC subpopulations in the same tumor. Thus, additional markers of functional stem cells are needed. The STAT3 pathway is a critical regulator of the function of normal stem cells, and evidence is accumulating for its important role in breast cancer stem cells. However, due to the lack of a method for separating live cells based on their level of STAT3 activity, it remains unknown whether STAT3 functions in the cancer stem cells themselves, or in surrounding niche cells, or in both. To approach this question, we constructed a series of lentiviral fluorescent (enhanced green fluorescent protein, EGFP) reporters that enabled flow cytometric enrichment of cells differing in STAT3-mediated transcriptional activity, as well as in vivo/in situ localization of STAT3 responsive cells. Using in vivo claudin-low cell line xenograft models of human breast cancer, we found that STAT3 signaling reporter activity (EGFP(+) ) is associated with a subpopulation of cancer cells enriched for mammosphere-forming efficiency, as well as TIC function in limiting dilution transplantation assays compared to negative or unsorted populations. Our results support STAT3 signaling activity as another functional marker for human breast cancer stem cells thus making it an attractive therapeutic target for stem-cell-directed therapy in some breast cancer subtypes.
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Affiliation(s)
- Wei Wei
- Lester & Sue Smith Breast Center, Baylor College of Medicine, Houston, Texas, USA; Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas, USA
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16
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Wedeking T, Löchte S, Richter CP, Bhagawati M, Piehler J, You C. Single Cell GFP-Trap Reveals Stoichiometry and Dynamics of Cytosolic Protein Complexes. NANO LETTERS 2015; 15:3610-3615. [PMID: 25901412 DOI: 10.1021/acs.nanolett.5b01153] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/04/2023]
Abstract
We developed in situ single cell pull-down (SiCPull) of GFP-tagged protein complexes based on micropatterned functionalized surface architectures. Cells cultured on these supports are lysed by mild detergents and protein complexes captured to the surface are probed in situ by total internal reflection fluorescence microscopy. Using SiCPull, we quantitatively mapped the lifetimes of various signal transducer and activator of transcription complexes by monitoring dissociation from the surface and defined their stoichiometry on the single molecule level.
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Affiliation(s)
- Tim Wedeking
- Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany
| | - Sara Löchte
- Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany
| | | | - Maniraj Bhagawati
- Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany
| | - Jacob Piehler
- Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany
| | - Changjiang You
- Department of Biology, University of Osnabrück, 49076 Osnabrück, Germany
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17
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Koirala S, Thomas LN, Too CKL. Prolactin/Stat5 and androgen R1881 coactivate carboxypeptidase-D gene in breast cancer cells. Mol Endocrinol 2014; 28:331-43. [PMID: 24433040 DOI: 10.1210/me.2013-1202] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
Plasma membrane-bound carboxypeptidase-D (CPD) cleaves C-terminal arginine from extracellular substrates. In the cell, arginine is converted to nitric oxide (NO). We have reported that up-regulation of CPD mRNA/protein levels by 17β-estradiol and prolactin (PRL) in breast cancer cells, and by testosterone in prostate cancer cells, increased NO production and cell survival. The CPD promoter contains a consensus γ-interferon-activated sequence (GAS) and 3 putative androgen response elements (ARE.1, ARE.2, ARE.3) that could potentially bind PRL-activated transcription factor Stat5 (signal transducer and activator of transcription 5) and the liganded androgen receptor (AR), respectively. This study showed that synthetic androgen R1881 and PRL elevated CPD mRNA/protein levels in human MCF-7 and T47D breast cancer cells in a time-/dose-dependent manner. PRL/R1881-elevated CPD expression was blocked by actinomycin-D, and a CPD promoter construct containing these GAS and AREs was stimulated by PRL or R1881, indicating transcriptional regulation by both hormones. Luciferase reporter assays showed that GAS and the adjacent ARE.1 only were active. Mutation of GAS in the ΔGAS-CPD construct (ARE.1 intact) abolished CPD promoter activity in response to PRL and, surprisingly, to R1881 as well. ΔGAS-CPD promoter activity was restored by PRL+R1881 in combination, and enhanced by ectopic Stat5, but abolished by Stat5 gene knockdown. Chromatin immunoprecipitation analysis confirmed binding of activated Stat5 and liganded AR to GAS and ARE.1, respectively. Activated Stat5 also induced binding of unliganded AR to ARE.1, and liganded AR induced binding of unactivated Stat5 to GAS. In summary, PRL and R1881, acting through Stat5 and AR, act cooperatively to stimulate CPD gene transcription in breast cancer cells.
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Affiliation(s)
- Samir Koirala
- Department of Biochemistry & Molecular Biology (S.K., L.N.T., C.K.L.T.) and Department of Obstetrics & Gynaecology (C.K.L.T.), Faculty of Medicine, Dalhousie University, Halifax, Nova Scotia B3H 4R2, Canada
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Identification of the kinase that activates a nonmetazoan STAT gives insights into the evolution of phosphotyrosine-SH2 domain signaling. Proc Natl Acad Sci U S A 2012; 109:E1931-7. [PMID: 22699506 DOI: 10.1073/pnas.1202715109] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/27/2023] Open
Abstract
SH2 domains are integral to many animal signaling pathways. By interacting with specific phosphotyrosine residues, they provide regulatable protein-protein interaction domains. Dictyostelium is the only nonmetazoan with functionally characterized SH2 domains, but the cognate tyrosine kinases are unknown. There are no orthologs of the animal tyrosine kinases, but there are very many tyrosine kinase-like kinases (TKLs), a group of kinases which, despite their family name, are classified mainly as serine-threonine kinases. STATs are transcription factors that dimerize via phosphotyrosine-SH2 domain interactions. STATc is activated by phosphorylation on Tyr922 when cells are exposed to the prestalk inducer differentiation inducing factor (DIF-1), a chlorinated hexaphenone. We show that in a null mutant for Pyk2, a tyrosine-specific TKL, exposure to DIF-1 does not activate STATc. Conversely, overexpression of Pyk2 causes constitutive STATc activation. Pyk2 phosphorylates STATc on Tyr922 in vitro and complexes with STATc both in vitro and in vivo. This demonstration that a TKL directly activates a STAT has significant implications for understanding the evolutionary origins of SH2 domain-phosphotyrosine signaling. It also has mechanistic implications. Our previous work suggested that a predicted constitutive STATc tyrosine kinase activity is counterbalanced in vivo by the DIF-1-regulated activity of PTP3, a Tyr922 phosphatase. Here we show that the STATc-Pyk2 complex is formed constitutively by an interaction between the STATc SH2 domain and phosphotyrosine residues on Pyk2 that are generated by autophosphorylation. Also, as predicted, Pyk2 is constitutively active as a STATc kinase. This observation provides further evidence for this highly atypical, possibly ancestral, STAT regulation mechanism.
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Shah S, Henriksen MA. A novel disrupter of telomere silencing 1-like (DOT1L) interaction is required for signal transducer and activator of transcription 1 (STAT1)-activated gene expression. J Biol Chem 2011; 286:41195-41204. [PMID: 22002246 PMCID: PMC3308833 DOI: 10.1074/jbc.m111.284190] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2011] [Revised: 09/21/2011] [Indexed: 01/14/2023] Open
Abstract
JAK-STAT-activated gene expression is both rapid and transient and requires dynamic post-translational modification of the chromatin template. Previously, we showed that following IFN-γ treatment, trimethylation of histone H3 at lysine 79 (H3K79me3) is rapidly and highly induced in the 5'-end of the STAT1-dependent gene interferon regulatory factor 1 (IRF1), but the role of this histone modification was unexplored. Here we report that DOT1L, the non-SET domain containing methyltransferase that modifies Lys-79, is localized across IRF1 in the uninduced state and is not further recruited by IFN-γ induction. RNAi-mediated depletion of DOT1L prevents the induction of H3K79me3 and lowers the transcription of IRF1 2-fold, as expected. Surprisingly, STAT1 binding to its DNA recognition element near the IRF1 promoter is diminished 2-fold in the DOT1L-depleted cell line. In vivo and in vitro protein interaction assays reveal a DOT1L-STAT1 interaction. Domain mapping identifies the middle region of DOT1L (amino acids 580-1183) as the STAT1 interaction domain. Overexpression of the DOT1L STAT1 interaction domain represses IRF1 transcription (2-fold) and interferes with STAT1 DNA binding at IRF1 and endogenous DOT1L histone methyltransferase activity. Collectively, our findings reveal a novel STAT1-DOT1L interaction that is required for the regulation JAK-STAT-inducible gene expression.
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Affiliation(s)
- Shaili Shah
- Department of Biology, The University of Virginia, Charlottesville, Virginia 22903
| | - Melissa A Henriksen
- Department of Biology, The University of Virginia, Charlottesville, Virginia 22903.
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Mohr A, Chatain N, Domoszlai T, Rinis N, Sommerauer M, Vogt M, Müller-Newen G. Dynamics and non-canonical aspects of JAK/STAT signalling. Eur J Cell Biol 2011; 91:524-32. [PMID: 22018664 DOI: 10.1016/j.ejcb.2011.09.005] [Citation(s) in RCA: 74] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2011] [Revised: 09/05/2011] [Accepted: 09/12/2011] [Indexed: 11/25/2022] Open
Abstract
The Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway directly links ligand-binding to a membrane-bound receptor with the activation of a transcription factor. This signalling module enables the cell to rapidly initiate a transcriptional response to external stimulation. The main components of this evolutionary conserved module are cytokines that specifically bind to cytokine receptors leading to the activation of receptor-associated Janus tyrosine kinases (JAKs). The receptor-bound JAKs activate STAT transcription factors through phosphorylation of a single tyrosine residue. Activated STAT dimers translocate into the nucleus to induce target gene expression. In this article we will review current opinions on the molecular mechanism and on intracellular dynamics of JAK/STAT signalling with a special focus on the cytokine receptor glycoprotein 130 (gp130) and STAT3. In particular we will concentrate on non-canonical aspects of Jak/STAT signalling including preassembled receptor complexes, preformed STAT dimers, STAT trafficking and non-canonical functions of STATs.
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Affiliation(s)
- Anne Mohr
- Institut für Biochemie und Molekularbiologie, RWTH Aachen University, Aachen, Germany
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21
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Qian CJ, Yao J, Si JM. Nuclear JAK2: form and function in cancer. Anat Rec (Hoboken) 2011; 294:1446-59. [PMID: 21809458 DOI: 10.1002/ar.21443] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2011] [Accepted: 05/19/2011] [Indexed: 12/23/2022]
Abstract
The conventional view of Janus kinase 2 (JAK2) is a nonreceptor tyrosine kinase which transmits information to the nucleus via the signal transducer and activator of transcriptions (STATs) without leaving the cytoplasm. However, accumulating data suggest that JAK2 may signal by exporting from cytoplasm to nucleus, where it guides the transcriptional machinery independent of STATs protein. Recent studies demonstrated that JAK2 is a crucial component of signaling pathways operating in the nucleus. Especially the latest landmark discovery confirmed that JAK2 goes into the nucleus and directly interacts with nucleoproteins, such as histone H3 at tyrosine 41 (H3Y41), nuclear factor 1-C2 (NF1-C2) and SWI/SNF-related helicases/ATPases (RUSH)-1α, indicating that JAK2 has a fresh nuclear function. Nuclear JAK2 is linked to a variety of cellular functions, such as cell cycle progression, apoptosis and genetic instability. The balance between these functions is an essential factor in determining whether a cell remains benign or becomes malignant. The aim of this review is intended to summarize the state of our knowledge on nuclear localization of JAK2 and nuclear JAK2 pathways, and to highlight the emerging roles for nuclear JAK2 in carcinogenesis.
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Affiliation(s)
- Cui-Juan Qian
- Department of Gastroenterology, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
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22
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Ge J, Wu H, Yao SQ. An unnatural amino acid that mimics phosphotyrosine. Chem Commun (Camb) 2010; 46:2980-2. [DOI: 10.1039/c000283f] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
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23
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Jerke U, Tkachuk S, Kiyan J, Stepanova V, Kusch A, Hinz M, Dietz R, Haller H, Fuhrman B, Dumler I. Stat1 nuclear translocation by nucleolin upon monocyte differentiation. PLoS One 2009; 4:e8302. [PMID: 20011528 PMCID: PMC2788426 DOI: 10.1371/journal.pone.0008302] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2009] [Accepted: 11/19/2009] [Indexed: 01/14/2023] Open
Abstract
BACKGROUND Members of the signal transducer and activator of transcription (Stat) family of transcription factors traverse the nuclear membrane through a specialized structure, called the nuclear pore complex (NPC), which represents a selective filter for the import of proteins. Karyophilic molecules can bind directly to a subset of proteins of the NPC, collectively called nucleoporins. Alternatively, the transport is mediated via a carrier molecule belonging to the importin/karyopherin superfamily, which transmits the import into the nucleus through the NPC. METHODOLOGY/PRINCIPAL FINDINGS In this study, we provide evidence for an alternative Stat1 nuclear import mechanism, which is mediated by the shuttle protein nucleolin. We observed Stat1-nucleolin association, nuclear translocation and specific binding to the regulatory DNA element GAS. Using expression of nucleolin transgenes, we found that the nuclear localization signal (NLS) of nucleolin is responsible for Stat1 nuclear translocation. We show that this mechanism is utilized upon differentiation of myeloid cells and is specific for the differentiation step from monocytes to macrophages. CONCLUSIONS/SIGNIFICANCE Our data add the nucleolin-Stat1 complex as a novel functional partner for the cell differentiation program, which is uniquely poised to regulate the transcription machinery via Stat1 and nuclear metabolism via nucleolin.
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Affiliation(s)
- Uwe Jerke
- Hannover Medical School, Hannover, Germany.
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Expression of JAKs/STATs pathway molecules in rat model of rapid focal segmental glomerulosclerosis. Pediatr Nephrol 2009; 24:1661-71. [PMID: 19350281 DOI: 10.1007/s00467-009-1163-4] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/12/2008] [Revised: 02/19/2009] [Accepted: 02/19/2009] [Indexed: 12/29/2022]
Abstract
The objective of this study was to investigate the role of the Janus kinase-signal transducers and activators of transcription (JAKs/STATs) pathway in focal segmental glomerulosclerosis. Sixty specific pathogen-free male Wistar rats were randomly divided into two groups: a model group (MG) and a control group (CG). In the MG group, nephropathy was induced by unilateral nephrectomy and a single tail vein injection of adriamycin (5 mg/kg). Ten rats were sacrificed every 2 weeks in each group. The expressions of smooth muscle alpha actin (alpha-SMA), collagen (COL)-IV, STAT1, and STAT3 were examined using histochemical techniques, and Western blotting was used to examine the protein levels of STAT1, STAT3, phosphorylated (P)-STAT1, P-STAT3, and transforming growth factor beta1 (TGFbeta(1)). The expressions of JAK1, JAK2, STAT1, STAT3, suppressors of cytokine signaling (SOCS)1, SOCS3, protein inhibitors of activated STAT (PIAS)1, and PIAS3 were also measured by real-time quantitative reverse transcriptase-PCR. A steady and significant increase in the expressions of alpha-SMA, COL-IV and TGFbeta(1) were observed in MG rats over the whole experimental course. Increased STAT1 and P-STAT1 levels in MG rats were observed by week 6, whereas increased levels of STAT3 and P-STAT3 were noted by week 2. At the mRNA levels, JAK1, STAT1, and PIAS1 were significantly increased in MG rats in week 2, whereas JAK2 mRNA showed a significant decrease by weeks 2 and 4, followed by an significant increase in week 6. Significantly increased STAT3 levels were noted in week 2, followed by a steady and significant decrease in weeks 4 and 6. Significantly reduced levels of SOCS1, SOCS3, and PIAS3 mRNA were noted at all time points. We conclude that the JAKs/STATs signaling pathway may play an important role in the pathological process of rapid focal segmental glomerulosclerosis in the rat model.
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Collet B, Ganne G, Bird S, Collins CM. Isolation and expression profile of a gene encoding for the Signal Transducer and Activator of Transcription STAT2 in Atlantic salmon (Salmo salar). DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY 2009; 33:821-829. [PMID: 19428483 DOI: 10.1016/j.dci.2009.01.007] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/20/2008] [Revised: 01/21/2009] [Accepted: 01/25/2009] [Indexed: 05/27/2023]
Abstract
Signal Transducer and Activator of Transcription (STAT)-2 is a molecule involved in the type I interferon (IFN) signalling pathway. The full length cDNA sequence of Atlantic salmon (Salmo salar) ssSTAT2 was determined and phylogenetic analysis of the amino acid sequence grouped this novel salmon gene to the STAT2 clade. This represents the first fish STAT2 report. The gene encodes for a 802 aa polypeptide that has 38% identity to the human or murine STAT2. The expression was monitored by qPCR in the kidney of animals over the time of infection with the Infectious Salmon Anaemia Virus (ISAV) and in TO cells infected with Infectious Pancreatic Necrosis Virus (IPNV) or with the Salmon Alphavirus (SAV). SAV and ISAV induced an approximate 10-fold increase in the level of expression of ssSTAT2 gene whilst IPNV only induced a 1.5-fold increase.
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Zhou X, Agazie YM. Molecular mechanism for SHP2 in promoting HER2-induced signaling and transformation. J Biol Chem 2009; 284:12226-34. [PMID: 19261604 DOI: 10.1074/jbc.m900020200] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
The Src homology phosphotyrosyl phosphatase 2 (SHP2) plays a positive role in HER2-induced signaling and transformation, but its mechanism of action is poorly understood. Given the significance of HER2 in breast cancer, defining a mechanism for SHP2 in the HER2 signaling pathway is of paramount importance. In the current report we show that SHP2 positively modulates the Ras-extracellular signal-regulated kinase 1 and 2 and the phospoinositide-3-kinase-Akt pathways downstream of HER2 by increasing the half-life the activated form of Ras. This is accomplished by dephosphorylating an autophosphorylation site on HER2 that serves as a docking platform for the SH2 domains of the Ras GTPase-activating protein (RasGAP). The net effect is an increase in the intensity and duration of GTP-Ras levels with the overall impact of enhanced HER2 signaling and cell transformation. In conformity to these findings, the HER2 mutant that lacks the SHP2 target site exhibits an enhanced signaling and cell transformation potential. Therefore, SHP2 promotes HER2-induced signaling and transformation at least in part by dephosphorylating a negative regulatory autophosphorylation site. These results suggest that SHP2 might serve as a therapeutic target against breast cancer and other cancers characterized by HER2 overexpression.
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Affiliation(s)
- Xiangdong Zhou
- Department of Biochemistry and The Marry Babb Randolph Cancer Center, West Virginia University, Morgantown, West Virginia 26506, USA
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Bouhet S, Lafont V, Billard E, Gross A, Dornand J. The IFNgamma-induced STAT1-CBP/P300 association, required for a normal response to the cytokine, is disrupted in Brucella-infected macrophages. Microb Pathog 2008; 46:88-97. [PMID: 19041714 DOI: 10.1016/j.micpath.2008.10.011] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2008] [Revised: 10/24/2008] [Accepted: 10/31/2008] [Indexed: 01/18/2023]
Abstract
To develop intracellularly within phagocytes and cause chronic infection, Brucella must overcome different steps of the host immune responses. IFNgamma is a key mediator of the innate and adaptive responses produced during Brucella infection. Therefore, Brucella would control host defenses by impairing macrophage responses to IFNgamma. We first showed that in infected human macrophages (VD3-differentiated THP-1 cells) Brucella escaped the microbicidal environment generated by IFNgamma. We then analyzed the IFNgamma-mediated signaling in Brucella-infected cells. We observed no decrease in STAT1 tyrosine or serine phosphorylation, or in dimerization of phosphorylated STAT1 (P-STAT1) and P-STAT1 translocation to the nucleus or in P-STAT1 binding to GAS, a minimal IFNgamma-response DNA sequence. In contrast, immuno-precipitation experiments indicated that the IFNgamma-mediated association of P-STAT1 with CBP/P300 transactivators was markedly reduced in infected macrophages, demonstrating that P-STAT1 was unable to normally recruit these transactivators. The host cell cAMP pathway triggered by Brucella could be responsible for this defect, CBP/P300 mobilization by phosphorylated CREB (P-CREB) disrupting the IFNgamma-induced STAT1-CBP/P300 association, required for a normal response of macrophages to IFNgamma. In any case, the inhibition of an essential protein-protein interaction probably lead to a deteriorated response to IFNgamma and thus participated in the pathogen's establishment within its host.
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Affiliation(s)
- Sandrine Bouhet
- Université Montpellier1, Centre d'étude d'Agents Pathogènes et Biotechnologies pour la Santé (CPBS), France
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Liu X, Ye L, Bai Y, Mojidi H, Simister NE, Zhu X. Activation of the JAK/STAT-1 signaling pathway by IFN-gamma can down-regulate functional expression of the MHC class I-related neonatal Fc receptor for IgG. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2008; 181:449-63. [PMID: 18566411 PMCID: PMC2667120 DOI: 10.4049/jimmunol.181.1.449] [Citation(s) in RCA: 68] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
Expression of many MHC genes is enhanced at the transcriptional or posttranscriptional level following exposure to the cytokine IFN-gamma. However, in this study we found that IFN-gamma down-regulated the constitutive expression of the neonatal Fc receptor (FcRn), an MHC class I-related molecule that functions to transport maternal IgG and protect IgG and albumin from degradation. Epithelial cell, macrophage-like THP-1 cell, and freshly isolated human PBMC exposure to IFN-gamma resulted in a significant decrease of FcRn expression as assessed by real-time RT-PCR and Western blotting. The down-regulation of FcRn was not caused by apoptosis or the instability of FcRn mRNA. Chromatin immunoprecipitation and gel mobility shift assays showed that STAT-1 bound to an IFN-gamma activation site in the human FcRn promoter region. Luciferase expression from an FcRn promoter-luciferase reporter gene construct was not altered in JAK1- and STAT-1-deficient cells following exposure to IFN-gamma, whereas expression of JAK1 or STAT-1 protein restored the IFN-gamma inhibitory effect on luciferase activity. The repressive effect of IFN-gamma on the FcRn promoter was selectively reversed or blocked by mutations of the core nucleotides in the IFN-gamma activation site sequence and by overexpression of the STAT-1 inhibitor PIAS1 or the dominant negative phospho-STAT-1 mutations at Tyr-701 and/or Ser-727 residues. Furthermore, STAT-1 might down-regulate FcRn transcription through sequestering the transcriptional coactivator CREB binding protein/p300. Functionally, IFN-gamma stimulation dampened bidirectional transport of IgG across a polarized Calu-3 lung epithelial monolayer. Taken together, our results indicate that the JAK/STAT-1 signaling pathway was necessary and sufficient to mediate the down-regulation of FcRn gene expression by IFN-gamma.
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Affiliation(s)
- Xindong Liu
- Laboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, MD 20742
| | - Lilin Ye
- Laboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, MD 20742
| | - Yu Bai
- Laboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, MD 20742
| | - Habi Mojidi
- Laboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, MD 20742
- Maryland Pathogen Research Institute, Graduate Program in Molecular and Cell Biology, University of Maryland, College Park, MD 20742
| | - Neil E. Simister
- Rosenstiel Center for Basic Biomedical Sciences and Biology Department, Brandeis University, Waltham, MA 02254
| | - Xiaoping Zhu
- Laboratory of Immunology, Virginia-Maryland Regional College of Veterinary Medicine, University of Maryland, College Park, MD 20742
- Maryland Pathogen Research Institute, Graduate Program in Molecular and Cell Biology, University of Maryland, College Park, MD 20742
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Transcription Factors STAT5 and STAT3. Prostate Cancer 2008. [DOI: 10.1007/978-1-60327-079-3_12] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022] Open
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Robertson G, Hirst M, Bainbridge M, Bilenky M, Zhao Y, Zeng T, Euskirchen G, Bernier B, Varhol R, Delaney A, Thiessen N, Griffith OL, He A, Marra M, Snyder M, Jones S. Genome-wide profiles of STAT1 DNA association using chromatin immunoprecipitation and massively parallel sequencing. Nat Methods 2007; 4:651-7. [PMID: 17558387 DOI: 10.1038/nmeth1068] [Citation(s) in RCA: 1041] [Impact Index Per Article: 57.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2007] [Accepted: 06/05/2007] [Indexed: 02/06/2023]
Abstract
We developed a method, ChIP-sequencing (ChIP-seq), combining chromatin immunoprecipitation (ChIP) and massively parallel sequencing to identify mammalian DNA sequences bound by transcription factors in vivo. We used ChIP-seq to map STAT1 targets in interferon-gamma (IFN-gamma)-stimulated and unstimulated human HeLa S3 cells, and compared the method's performance to ChIP-PCR and to ChIP-chip for four chromosomes. By ChIP-seq, using 15.1 and 12.9 million uniquely mapped sequence reads, and an estimated false discovery rate of less than 0.001, we identified 41,582 and 11,004 putative STAT1-binding regions in stimulated and unstimulated cells, respectively. Of the 34 loci known to contain STAT1 interferon-responsive binding sites, ChIP-seq found 24 (71%). ChIP-seq targets were enriched in sequences similar to known STAT1 binding motifs. Comparisons with two ChIP-PCR data sets suggested that ChIP-seq sensitivity was between 70% and 92% and specificity was at least 95%.
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Affiliation(s)
- Gordon Robertson
- British Columbia Cancer Agency Genome Sciences Centre, 675 West 10th Avenue, Vancouver, British Columbia V5Z 4S6, Canada
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Ganster RW, Guo Z, Shao L, Geller DA. Differential effects of TNF-alpha and IFN-gamma on gene transcription mediated by NF-kappaB-Stat1 interactions. J Interferon Cytokine Res 2006; 25:707-19. [PMID: 16318585 DOI: 10.1089/jir.2005.25.707] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
Regulation of gene transcription by the cytokines tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) involves complex interactions between NF-kappaB and Stat families of transcription factors. The purpose of this study was to identify the spatial promoter requirements that govern cytokine synergy for gene transcription regulated by NF-kappaB and Stat factors. Using a set of transcription reporter-luciferase constructs, we show that the relative orientation of juxtaposed NF-kappaB-Stat (SIE) cis-elements determines the ability of TNF-alpha and IFN- gamma to induce gene transcription. Further, NF-kappaB and Stat1 proteins directly regulate transcription by interacting cooperatively on NF-kappaB-SIE DNA binding in response to TNF-alpha plus IFN-gamma. Coimmunoprecipitation provides evidence for a direct NF-kappaB/Stat1 protein-protein interaction. In contrast, IFN-gamma inhibits TNF-alpha-induced transcription of an NF-kappaB reporter gene in a Stat1-dependent mechanism in 2fTGH fibroblasts. Similarly, Stat1 is inhibitory to NF-kappaB overexpression-induced transcription. IFN-gamma and Stat1-dependent inhibition of NF-kappaB transcription occurs independent of TNF-alpha-induced NF-kappaB DNA binding. Interestingly, IFN-gamma pretreatment of 2fTGH fibroblasts potentiates TNF-alpha induction of Stat1 DNA binding. Further, ChIP analysis was applied to detect cytokine-induced in vivo binding and transcriptional regulation of the human inducible nitric oxide synthase (iNOS) gene by NF-kappaB and Stat1. These data demonstrate complex transcriptional regulatory mechanisms elicited by TNF-alpha and IFN-gamma and have potentially important implications for other genes differentially controlled by cytokines.
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Affiliation(s)
- Raymond W Ganster
- Department of Surgery, University of Pittsburgh School of Medicine, PA 15261, USA
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Gartsbein M, Alt A, Hashimoto K, Nakajima K, Kuroki T, Tennenbaum T. The role of protein kinase C δ activation and STAT3 Ser727 phosphorylation in insulin-induced keratinocyte proliferation. J Cell Sci 2006; 119:470-81. [PMID: 16418226 DOI: 10.1242/jcs.02744] [Citation(s) in RCA: 54] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023] Open
Abstract
Activation of the STAT family of transcription factors is regulated by cytokines and growth factors. STAT tyrosine and serine phosphorylation are linked to the transcriptional activation and function of STAT. We have previously described a unique pathway inducing keratinocyte proliferation, which is mediated by insulin stimulation and depends on protein kinase C δ (PKCδ). In this study, we assessed STAT3 activation downstream of this pathway and characterized the role of PKCδ activation in STAT3 tyrosine and serine phosphorylation and keratinocyte proliferation. Following insulin stimulation, STAT3 interacted with PKCδ but not with any other PKC isoform expressed in skin. Activated forms of PKCδ and STAT3 were essential for insulin-induced PKCδ-STAT3 activation in keratinocyte proliferation. Abrogation of PKCδ activity inhibited insulin-induced STAT3 phosphorylation, PKCδ-STAT3 association and nuclear translocation. In addition, overexpression of STAT3 tyrosine mutant eliminated insulin-induced PKCδ activation and keratinocyte proliferation. Finally, overexpression of a STAT3 serine mutant abrogated insulin-induced STAT3 serine phosphorylation and STAT3-induced keratinocyte proliferation, whereas STAT3 tyrosine phosphorylation was induced and nuclear localization remained intact. This study indicates that PKCδ activation is a primary regulator of STAT3 serine phosphorylation and that PKCδ is essential in directing insulin-induced signaling in keratinocyte proliferation.
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Affiliation(s)
- Marina Gartsbein
- Faculty of Life Sciences, Bar Ilan University, Ramat-Gan, 52900 Israel
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Shushakova N, Tkachuk N, Dangers M, Tkachuk S, Park JK, Zwirner J, Hashimoto K, Haller H, Dumler I. Urokinase-induced activation of the gp130/Tyk2/Stat3 pathway mediates a pro-inflammatory effect in human mesangial cells via expression of the anaphylatoxin C5a receptor. J Cell Sci 2005; 118:2743-53. [PMID: 15944400 DOI: 10.1242/jcs.02409] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Glomerular mesangial cells (MCs) are central to the pathogenesis of progressive glomeruli-associated renal diseases. However, molecular mechanisms underlying changes in MC functions still remain poorly understood. Here, we show that in MCs, the urokinase-type plasminogen activator (uPA) induces, via its specific receptor (uPAR, CD87), upregulated expression of the complement anaphylatoxin C5a receptor (C5aR, CD88), and modulates C5a-dependent functional responses. This effect is mediated via the interaction of the uPA-specific receptor (uPAR, CD87) and gp130, a signal transducing subunit of the receptor complexes for the IL-6 cytokine family. The Janus kinase Tyk2 and the transcription factor Stat3 serve as downstream components in the signaling cascade resulting in upregulation of C5aR expression. In vivo, expression of C5aR and uPAR was increased in the mesangium of wild-type mice in a lipopolysaccharide (LPS)-induced model of inflammation, whereas in uPAR(-/-) animals C5aR expression remained unchanged. This is the first demonstration in vitro and in vivo that uPA acts in MCs as a modulator of immune responses via control of immune-competent receptors. The data suggest a novel role for uPA/uPAR in glomeruli-associated renal failure via a signaling cross-talk between the fibrinolytic and immune systems.
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Mann-Chandler MN, Kashyap M, Wright HV, Norozian F, Barnstein BO, Gingras S, Parganas E, Ryan JJ. IFN-gamma induces apoptosis in developing mast cells. THE JOURNAL OF IMMUNOLOGY 2005; 175:3000-5. [PMID: 16116187 DOI: 10.4049/jimmunol.175.5.3000] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Mast cells are critical effectors of allergic disease, and are now implicated in immune responses observed in arthritis, multiple sclerosis, and heart disease. Because of their role in inflammation, understanding how mast cells develop is of clinical importance. In this study we determined the effects of IFN-gamma on mast cell survival. Using in vitro culture of bone marrow cells in IL-3 plus stem cell factor, we found that the addition of IFN-gamma induced apoptosis, as exhibited by the presence of subdiploid DNA and caspase activation. IFN-gamma-mediated apoptosis was Stat1-dependent, and was accompanied by loss of mitochondrial membrane potential. Apoptosis was reduced in cultures of bone marrow cells derived from p53- or Bax-deficient mice, as well as H2K-Bcl-2 transgenic mice. IFN-gamma hyperresponsiveness has been shown to result in inflammatory disease and death in mice lacking the regulatory protein suppressor of cytokine signaling (SOCS)-1. Bone marrow cells from SOCS-1 knockout (KO) mice failed to give rise to viable mast cells after culture in IL-3 plus stem cell factor, with profound apoptosis occurring as the cultures matured. However, bone marrow cells lacking both SOCS-1 and IFN-gamma survived normally. This in vitro defect in mast cell development was recapitulated in vivo. SOCS-1 KO mice demonstrated a 67% decrease in peritoneal mast cell numbers relative to wild-type mice, a deficiency that was reversed in SOCS-1/IFN-gamma KO mice. These data demonstrate the potent regulatory effects of IFN-gamma on mast cell survival and show that this cytokine can elicit mast cell death in vitro and in vivo.
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Ma Z, Chang MJ, Shah RC, Benveniste EN. Interferon-gamma-activated STAT-1alpha suppresses MMP-9 gene transcription by sequestration of the coactivators CBP/p300. J Leukoc Biol 2005; 78:515-23. [PMID: 15894584 DOI: 10.1189/jlb.0205112] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
Interferon-gamma (IFN-gamma) is a pleiotropic cytokine involved in aspects of immune regulation, cell proliferation, and host defense mechanisms directed toward various cancers. Some of the biological functions of IFN-gamma are achieved through inhibition of gene expression, although the mechanisms by which IFN-gamma suppresses gene transcription are poorly understood. Herein, we demonstrate the molecular basis by which IFN-gamma mediates suppression of the matrix metalloproteinase-9 (MMP-9) gene. IFN-gamma-activated signal transducer and activator of transcription-1alpha (STAT-1alpha) suppresses MMP-9 gene transcription, which is dependent on phosphorylation of tyrosine 701 but not phosphorylation of serine 727. The coactivator cyclic AMP response element-binding protein-binding protein (CBP) is an important component of induction of MMP-9 gene transcription. IFN-gamma induces the in vivo association of STAT-1alpha and CBP and decreases the association of CBP to the MMP-9 promoter. IFN-gamma does not influence the stability of CBP nor does IFN-gamma affect chromatin-remodeling events on the MMP-9 promoter. IFN-gamma inhibits the assembly of the MMP-9 transcription complex by suppressing H3/H4 acetylation and inhibiting recruitment of Pol II to the MMP-9 promoter. These findings indicate that IFN-gamma/STAT-1alpha exert their inhibitory effects by affecting multiple aspects of MMP-9 gene transcription.
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Affiliation(s)
- Zhendong Ma
- Department of Cell Biology, University of Alabama at Birmingham, Birmingham, AL 35294-0005, USA
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Kambhampati S, Verma A, Li Y, Parmar S, Sassano A, Platanias LC. Signalling pathways activated by all-trans-retinoic acid in acute promyelocytic leukemia cells. Leuk Lymphoma 2005; 45:2175-85. [PMID: 15512805 DOI: 10.1080/10428190410001722053] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/26/2022]
Abstract
Acute promyelocytic leukemia is a form of acute myelogenous leukemia, characterized by the t(15;17) chromososmal translocation and the presence of the abnormal PML-RARalpha fusion protein. All-trans-retinoic acid is a potent agent for the treatment of this fatal subtype of AML, and is particularly effective when combined with cytotoxic chemotherapy. The important biological activities of all-trans-retinoic acid in vitro and in vivo have provoked extensive studies over the years, aimed to define the mechanisms by which it induces its antileukemic effects. It is now well established that all-trans-retinoic acid when administered at pharmacological doses can reverse the dominant-negative effects that the PML-RARalpha oncoprotein exhibits on the functions of the wild type PML and RARalpha proteins. All-trans-retinoic acid induces gene transcription via retinoic acid responsive elements (RARE) that are present in the promoters of retinoid-responsive genes that ultimately result in the production of protein products that regulate leukemic cell differentiation and induce cell-cycle arrest. There is now accumulating evidence that additional signalling pathways are activated during all-trans-retinoic acid-treatment of cells, involving Stat-proteins, tyrosine kinases and mitogen-activated protein (Map) kinases. This review summarizes the current knowledge on the signalling cascades activated by all-trans-retinoic acid in APL cells. The clinical implications and potential translational applications from the accumulating knowledge in the field are also discussed.
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Affiliation(s)
- Suman Kambhampati
- Robert H. Lurie Comprehensive Cancer Center and Division of Hematology-Oncology, Northwestern University Medical School, Chicago, IL 60611, USA
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Thomas M, Finnegan CE, Rogers KMA, Purcell JW, Trimble A, Johnston PG, Boland MP. STAT1: a modulator of chemotherapy-induced apoptosis. Cancer Res 2005; 64:8357-64. [PMID: 15548705 DOI: 10.1158/0008-5472.can-04-1864] [Citation(s) in RCA: 78] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
The anthracyclines, such as doxorubicin, are widely used in the treatment of breast cancer. Previously, we showed that these drugs could activate the transcription factor, nuclear factor kappaB, in a DNA damage-dependent manner. We now show that these drugs can potentiate the activation of signal transducer and activator of transcription 1 (STAT1) in MDA-MB 435 breast cancer cells treated with IFN-gamma. We observed that key markers of STAT1 activation, including tyrosine 701 and serine 727 phosphorylation, were enhanced in the presence of doxorubicin. This potentiation resulted in enhanced nuclear localization of activated STAT1 and led to an increase in the nuclear binding of activated STAT complexes. The observed potentiation was specific for STAT1 and IFN-gamma, as no effects were observed with either STAT3 or STAT5. Furthermore, the type I IFNs (alpha and beta) had little or no effect. The observed effects on STAT1 phosphorylation have previously been linked with maximal transcriptional activation and apoptosis. Cell viability was assessed by crystal violet staining followed by analysis with CalcuSyn to determine combination index values, a measure of synergy. We confirmed that significant synergy existed between IFN-gamma and doxorubicin (combination index = 0.34) at doses lower than IC(50) values for this drug (0.67 micromol/L). In support of this, we observed that apoptotic cell death was also enhanced by measuring poly(ADP-ribose) polymerase and caspase-3 cleavage. Finally, suppression of STAT1 expression by small-interfering RNA resulted in a loss of synergistic apoptotic cell death compared with cells, where no suppression of STAT1 expression was attained with scrambled small-interfering RNA control. We conclude that doxorubicin potentiates STAT1 activation in response to IFN-gamma, and that this combination results in enhanced apoptosis in breast cancer cells.
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Affiliation(s)
- Michelle Thomas
- Centre for Cancer Research, Queens University Belfast, Belfast City Hospital, Belfast, Northern Ireland
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Wiwi CA, Waxman DJ. Role of Hepatocyte Nuclear Factors in Transcriptional Regulation of Male-specific CYP2A2. J Biol Chem 2005; 280:3259-68. [PMID: 15539409 DOI: 10.1074/jbc.m409294200] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Cytochrome P450 2A2 (CYP2A2) is an adult male-specific rat liver steroid hydroxylase whose sex-dependent expression is regulated at the transcriptional level by sexually dimorphic pituitary growth hormone (GH) secretory patterns. In contrast to CYP2C11 and other male-specific, plasma GH pulse-inducible liver genes, CYP2A2 is highly expressed in hypophysectomized rat liver, despite the absence of GH stimulation. CYP2A2 promoter fragments 0.9-6.2 kb long exhibited unusually high basal promoter activity when transfected into the liver cell line HepG2. A further approximately 2.5-fold increase in activity was obtained by cotransfection of hepatocyte nuclear factor (HNF) 3gamma or HNF4alpha. CYP2A2 promoter activity was inhibited approximately 85% by transfection of HNF3beta or HNF6, both of which are more highly expressed in female than male liver and can strongly trans-activate the female-specific CYP2C12 promoter. The male GH pulse-activated transcription factor STAT5b had no effect on CYP2A2 promoter activity, either alone or in combination with HNF3gamma and HNF4alpha, consistent with the GH pulse-independence of CYP2A2 expression. By contrast, STAT5b synergistically enhanced the transcriptional activity of HNF4alpha toward two other male-specific liver target genes, Cyp2d9 and CYP8B1. Furthermore, STAT5b in combination with the HNF4alpha coactivator peroxisome proliferator-activated receptor gamma coactivator-1alpha strongly enhanced the transcriptional activity of HNF4alpha toward CYP8B1 but not toward CYP2A2. These findings support the hypothesis that sex-dependent HNFs contribute to the sexually dimorphic expression of CYP2A2 and other liver CYPs and highlight the ability of STAT5b to act in concert with HNF4alpha to regulate select male-specific liver CYP genes.
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Affiliation(s)
- Christopher A Wiwi
- Division of Cell and Molecular Biology, Department of Biology Boston University, Boston, Massachusetts 02215, USA
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Xia B, Yu YH, Guo QS, Li XY, Jiang L, Li J. Association of Fas-670 gene polymorphism with inflammatory bowel disease in Chinese patients. World J Gastroenterol 2005; 11:415-7. [PMID: 15637757 PMCID: PMC4205351 DOI: 10.3748/wjg.v11.i3.415] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: Recent studies suggest that Fas-mediated apoptosis is involved in the pathogenesis of inflammatory bowel disease (IBD). It has been hypothesized that either increased apoptosis of intestinal epithelium or decreased apoptosis of lamina propria lymphocytes may induce inflammation of gut. The aim of this study was to determine whether the Fas gene promoter polymorphism at position-670 was associated with IBD in Chinese patients.
METHODS: Fifty unrelated Chinese patients with IBD (38 patients with ulcerative colitis and 12 with Crohn’s disease) and 124 healthy controls were genotyped for the Fas-670 polymorphism by PCR-restriction fragment length polymorphism method. The PCR product was digested by Mva I restriction enzyme.
RESULTS: Distribution of the Fas-670 gene polymorphism was 33% for the AA genotype, 52% for the AG genotype and 15% for the GG genotype in 124 healthy subjects. In patients with IBD, 30% was for the AA genotype, 42% for the AG genotype and 28% for the GG genotype respectively. However, there was no significant difference in the genotype (P = 0.1498), allele frequencies (P = 0.3198) and carriage frequencies (P = 0.4133) between healthy controls and IBD patients. Furthermore, we did not find any difference between the left-sided colitis and total colitis (P = 0.8242).
CONCLUSION: Fas-670 polymorphism is not associated with IBD in Chinese patients.
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Affiliation(s)
- Bing Xia
- Department of Internal Medicine, Zhongnan Hospital, Medical School of Wuhan University, Wuhan 430071, Hubei Province, China.
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40
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DeVries TA, Kalkofen RL, Matassa AA, Reyland ME. Protein Kinase Cδ Regulates Apoptosis via Activation of STAT1. J Biol Chem 2004; 279:45603-12. [PMID: 15322115 DOI: 10.1074/jbc.m407448200] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022] Open
Abstract
Protein kinase Cdelta (PKCdelta) is required for mitochondria-dependent apoptosis; however, little is known about downstream effectors of PKCdelta in apoptotic cells. Here we show that activation of STAT1 is an early response to DNA damage and that STAT1 activation requires PKCdelta. Treatment of HeLa cells with etoposide results in phosphorylation of STAT1 on Ser(727) and the association of STAT1 with PKCdelta. Etoposide increases transcription from STAT1-dependent reporter constructs. Increased transcription, as well as STAT1 Ser(727) phosphorylation, can be blocked by inhibition or depletion of PKCdelta. To ask if STAT1 is required for PKCdelta-mediated apoptosis, we utilized U3A STAT1-deficient cells. Induction of apoptosis by PKCdelta is suppressed in U3A cells but can be rescued by co-transfection with STAT1alpha but not STAT1 mutated at Ser(727). Nuclear accumulation of STAT1, phospho-Ser(727) STAT1, and PKCdelta are detectable 30-60 min after treatment with etoposide. Nuclear localization is necessary for apoptosis, since a nuclear localization mutant of PKCdelta does not induce apoptosis in U3A cells reconstituted with STAT1alpha, and a nuclear localization mutant of STAT1 does not support PKCdelta-induced apoptosis in U3A cells. Our data identify STAT1 as a downstream target of PKCdelta and suggest that PKCdelta may regulate apoptosis by activation of STAT1 target genes.
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Affiliation(s)
- Tracie A DeVries
- Department of Craniofacial Biology, School of Dentistry, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA
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41
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Srivastava KK, Batra S, Sassano A, Li Y, Majchrzak B, Kiyokawa H, Altman A, Fish EN, Platanias LC. Engagement of Protein Kinase C-θ in Interferon Signaling in T-cells. J Biol Chem 2004; 279:29911-20. [PMID: 15150272 DOI: 10.1074/jbc.m401997200] [Citation(s) in RCA: 45] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Protein kinase C-theta (PKC-theta) plays important roles in the activation and survival of lymphocytes and is the predominant PKC isoform expressed in T-cells. Interferons regulate T-cell function and activation, but the precise signaling mechanisms by which they mediate such effects have not been elucidated. We determined whether PKC-theta is engaged in interferon (INF) signaling in T-cells. Both Type I (alpha, beta) and Type II (gamma) IFNs induced phosphorylation of PKC-theta in human T-cell lines and primary human T-lymphocytes. Such phosphorylation of PKC-theta resulted in activation of its kinase domain, suggesting that this kinase plays a functional role in interferon signaling. Consistent with this, inhibition of PKC-theta protein expression using small interfering RNAs (siRNA) abrogated IFN-alpha- and IFN-gamma-dependent gene transcription via GAS elements. Similarly, blocking of PKC-theta kinase activity by overexpression of a dominant-negative PKC-theta mutant also blocked GAS-driven transcription, further demonstrating a requirement for PKC-theta in IFN-dependent transcriptional activation. The effects of PKC-theta on IFN-dependent gene transcription were not mediated by regulation of the IFN-activated STAT pathway, as siRNA-mediated PKC-theta knockdown had no effects on STAT1 phosphorylation and binding of STAT1-containing complexes to SIE/GAS elements. On the other hand, siRNA-mediated PKC-theta inhibition blocked phosphorylation/activation of MKK4, suggesting that interferon-dependent PKC-theta activation regulates downstream engagement of MAP kinase pathways. Altogether, these findings demonstrate that PKC-theta is an interferon-inducible kinase and strongly suggest that it plays an important role in the generation of interferon-responses in T-cells.
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Affiliation(s)
- Kishore K Srivastava
- Robert H. Lurie Comprehensive Cancer Center and Division of Hematology-Oncology, Northwestern University Medical School and Lakeside Veterans Administration Medical Center, Chicago, Illinois 60611, USA
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42
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Sarcar B, Ghosh AK, Steele R, Ray R, Ray RB. Hepatitis C virus NS5A mediated STAT3 activation requires co-operation of Jak1 kinase. Virology 2004; 322:51-60. [PMID: 15063116 DOI: 10.1016/j.virol.2004.01.008] [Citation(s) in RCA: 46] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2003] [Revised: 12/09/2003] [Accepted: 01/05/2004] [Indexed: 12/21/2022]
Abstract
Hepatitis C virus (HCV) is a major etiologic agent for chronic hepatitis worldwide and often leads to cirrhosis and hepatocellular carcinoma. However, the mechanism for development of chronic hepatitis or hepatocarcinogenesis by HCV remains unclear. Signal transducers and activators of transcription (STATs) family proteins function as the downstream effectors of cytokine signaling and play a critical role in cell growth regulation. In many cancers including liver, STAT3 is often constitutively activated, although the mechanism of persistent activation of STAT3 is unknown. The nonstructural protein 5A (NS5A) encoded from the HCV genome has shown cell growth regulatory properties. In this study, we have observed that HCV NS5A activates STAT3 phosphorylation, which in turn translocates into the nucleus. In vivo activation of STAT3 was also observed in the liver of transgenic mice expressing HCV NS5A. Introduction of NS5A in hepatoma cells modulated STAT3 downstream molecules Bcl-xL and p21 expression. To determine if STAT3 activation by NS5A could induce STAT3 mediated gene expression, a luciferase reporter construct based on a synthetic promoter was used to transfect hepatoma cells. Activation of endogenous cellular STAT3 by HCV NS5A induced luciferase gene expression through STAT3 specific binding elements. Our subsequent studies suggested that NS5A forms a complex with Jak1 and recruits STAT3 for activation. Taken together, our results suggested that NS5A activates STAT3 through co-operation of Jak1 kinase and activated STAT3 may contribute to HCV-mediated pathogenesis.
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Affiliation(s)
- Bhaswati Sarcar
- Department of Pathology, Saint Louis University, St. Louis, MO 63104, USA
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43
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Zhukovskaya NV, Fukuzawa M, Tsujioka M, Jermyn KA, Kawata T, Abe T, Zvelebil M, Williams JG. Dd-STATb, a Dictyostelium STAT protein with a highly aberrant SH2 domain, functions as a regulator of gene expression during growth and early development. Development 2004; 131:447-58. [PMID: 14701681 DOI: 10.1242/dev.00927] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Dictyostelium, the only known non-metazoan organism to employ SH2 domain:phosphotyrosine signaling, possesses STATs (signal transducers and activators of transcription) and protein kinases with orthodox SH2 domains. Here, however, we describe a novel Dictyostelium STAT containing a remarkably divergent SH2 domain. Dd-STATb displays a 15 amino acid insertion in its SH2 domain and the conserved and essential arginine residue, which interacts with phosphotyrosine in all other known SH2 domains, is substituted by leucine. Despite these abnormalities, Dd-STATb is biologically functional. It has a subtle role in growth, so that Dd-STATb-null cells are gradually lost from the population when they are co-cultured with parental cells, and microarray analysis identified several genes that are either underexpressed or overexpressed in the Dd-STATb null strain. The best characterised of these,discoidin 1, is a marker of the growth-development transition and it is overexpressed during growth and early development of Dd-STATb null cells. Dimerisation of STAT proteins occurs by mutual SH2 domain:phosphotyrosine interactions and dimerisation triggers STAT nuclear accumulation. Despite its aberrant SH2 domain, the Dd-STATb protein sediments at the size expected for a homodimer and it is constitutively enriched in the nucleus. Moreover, these properties are retained when the predicted site of tyrosine phosphorylation is substituted by phenylalanine. These observations suggest a non-canonical mode of activation of Dd-STATb that does not rely on orthodox SH2 domain:phosphotyrosine interactions.
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Affiliation(s)
- Natasha V Zhukovskaya
- School of Life Sciences, University of Dundee, MSI/WTB Complex, Dow Street, Dundee DD1 5EH, UK
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44
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Li Y, Sassano A, Majchrzak B, Deb DK, Levy DE, Gaestel M, Nebreda AR, Fish EN, Platanias LC. Role of p38α Map Kinase in Type I Interferon Signaling. J Biol Chem 2004; 279:970-9. [PMID: 14578350 DOI: 10.1074/jbc.m309927200] [Citation(s) in RCA: 93] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023] Open
Abstract
Multiple signaling pathways are activated during engagement of the Type I interferon (IFN) receptor to mediate biological responses, including the Jak-Stat and Rac1/p38 Map kinase signaling cascades. In the present study we sought to determine the functional relevance of the p38alpha isoform in IFN signaling, using cells from mouse embryos with targeted disruption of the p38alpha gene. Our data demonstrate that p38alpha activation is essential for Type I IFN-dependent transcriptional regulation via ISRE or GAS elements. On the other hand, the function of p38alpha is not required for IFN-dependent Ser727 or Tyr701 phosphorylation of Stat1 and does not impact on the formation of ISGF3 or SIF nuclear binding complexes. In efforts to identify downstream effectors of p38 that may mediate IFN-dependent transcriptional responses, we found that IFNalpha activates the kinase Msk1, a known regulator of histone phosphorylation and chromatin remodeling. In other studies, we demonstrate that Type I IFN-dependent activation of the kinases MapKapK-2 and MapKapK-3 is defective in the absence of p38alpha, while Type I IFN-dependent antiviral properties are decreased in cells with targeted disruption of the MapKapK-2 gene. Altogether, our data establish that the p38alpha Map kinase pathway regulates activation of downstream effectors that participate in the induction of IFN-dependent gene transcription, to mediate IFN-responses.
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Affiliation(s)
- Yongzhong Li
- Robert H. Lurie Comprehensive Cancer Center, Northwestern University Medical School and Lakeside Veterans Administration Medical Center, Chicago, Illinois 60611, USA
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45
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Bolander FF. Hormonally Regulated Transcription Factors. Mol Endocrinol 2004. [DOI: 10.1016/b978-012111232-5/50013-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/23/2022] Open
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46
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Davoodi-Semiromi A, Laloraya M, Kumar GP, Purohit S, Jha RK, She JX. A mutant Stat5b with weaker DNA binding affinity defines a key defective pathway in nonobese diabetic mice. J Biol Chem 2003; 279:11553-61. [PMID: 14701862 DOI: 10.1074/jbc.m312110200] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022] Open
Abstract
A number of cytokines that finely regulate immune response have been implicated in the pathogenesis or protection of type 1 diabetes and other autoimmune diseases. It is, therefore, of pivotal importance to examine a family of proteins that serve as signal transducers and activators of transcription (STATs), which regulate the transcription of a variety of cytokines. We report here a defective gene (Stat5b) located on chromosome 11 within a previously mapped T1D susceptibility interval (Idd4) in the nonobese diabetic (NOD) mice. Our sequencing analysis revealed a unique mutation C1462A that results in a leucine to methionine (L327M) in Stat5b of NOD mice. Leu(327), the first residue in the DNA binding domain of STAT proteins, is conserved in all identified mammalian STAT proteins. Homology modeling predicted that the mutant Stat5b has a weaker DNA binding, which was confirmed by DNA-protein binding assays. The inapt transcriptional regulation ability of the mutated Stat5b is proved by decreased levels of RNA of Stat5b-regulated genes (IL-2Rbeta and Pim1). Consequently, IL-2Rbeta and Pim1 proteins were shown by Western blotting to have lower levels in NOD compared with normal B6 mice. These proteins have been implicated in immune regulation, apoptosis, activation-induced cell death, and control of autoimmunity. Therefore, the Stat5b pathway is a key molecular defect in NOD mice.
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Affiliation(s)
- Abdoreza Davoodi-Semiromi
- Department of Pathology, Immunology, and Laboratory Medicine, College of Medicine, University of Florida, Gainesville, Florida 32610, USA
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47
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Calò V, Migliavacca M, Bazan V, Macaluso M, Buscemi M, Gebbia N, Russo A. STAT proteins: from normal control of cellular events to tumorigenesis. J Cell Physiol 2003; 197:157-68. [PMID: 14502555 DOI: 10.1002/jcp.10364] [Citation(s) in RCA: 461] [Impact Index Per Article: 21.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/20/2023]
Abstract
Signal transducers and activators of transcription (STAT) proteins comprise a family of transcription factors latent in the cytoplasm that participate in normal cellular events, such as differentiation, proliferation, cell survival, apoptosis, and angiogenesis following cytokine, growth factor, and hormone signaling. STATs are activated by tyrosine phosphorylation, which is normally a transient and tightly regulates process. Nevertheless, several constitutively activated STATs have been observed in a wide number of human cancer cell lines and primary tumors, including blood malignancies and solid neoplasias. STATs can be divided into two groups according to their specific functions. One is made up of STAT2, STAT4, and STAT6, which are activated by a small number of cytokines and play a distinct role in the development of T-cells and in IFNgamma signaling. The other group includes STAT1, STAT3, and STAT5, activated in different tissues by means of a series of ligands and involved in IFN signaling, development of the mammary gland, response to GH, and embriogenesis. This latter group of STATS plays an important role in controlling cell-cycle progression and apoptosis and thus contributes to oncogenesis. Although an increased expression of STAT1 has been observed in many human neoplasias, this molecule can be considered a potential tumor suppressor, since it plays an important role in growth arrest and in promoting apoptosis. On the other hand, STAT3 and 5 are considered as oncogenes, since they bring about the activation of cyclin D1, c-Myc, and bcl-xl expression, and are involved in promoting cell-cycle progression, cellular transformation, and in preventing apoptosis.
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Affiliation(s)
- Valentina Calò
- Section of Molecular Oncology, Department of Oncology, Regional Reference Center for the Biomolecular Characterization of Neoplasms and Genetic Screening of Hereditary Tumors, University of Palermo, Palermo, Italy
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48
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Agazie YM, Hayman MJ. Molecular mechanism for a role of SHP2 in epidermal growth factor receptor signaling. Mol Cell Biol 2003; 23:7875-86. [PMID: 14560030 PMCID: PMC207628 DOI: 10.1128/mcb.23.21.7875-7886.2003] [Citation(s) in RCA: 239] [Impact Index Per Article: 10.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/19/2003] [Revised: 06/16/2003] [Accepted: 07/29/2003] [Indexed: 11/20/2022] Open
Abstract
The Src homology 2-containing phosphotyrosine phosphatase (SHP2) is primarily a positive effector of receptor tyrosine kinase signaling. However, the molecular mechanism by which SHP2 effects its biological function is unknown. In this report, we provide evidence that defines the molecular mechanism and site of action of SHP2 in the epidermal growth factor-induced mitogenic pathway. We demonstrate that SHP2 acts upstream of Ras and functions by increasing the half-life of activated Ras (GTP-Ras) in the cell by interfering with the process of Ras inactivation catalyzed by Ras GTPase-activating protein (RasGAP). It does so by inhibition of tyrosine phosphorylation-dependent translocation of RasGAP to the plasma membrane, to its substrate (GTP-Ras) microdomain. Inhibition is achieved through the dephosphorylation of RasGAP binding sites at the level of the plasma membrane. We have identified Tyr992 of the epidermal growth factor receptor (EGFR) to be one such site, since its mutation to Phe renders the EGFR refractory to the effect of dominant-negative SHP2. To our knowledge, this is the first report to outline the site and molecular mechanism of action of SHP2 in EGFR signaling, which may also serve as a model to describe its role in other receptor tyrosine kinase signaling pathways.
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Affiliation(s)
- Yehenew M Agazie
- Department of Molecular Genetics and Microbiology, Health Sciences Center, Stony Brook University, Stony Brook, New York 11794-5222, USA
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49
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Schroder K, Hertzog PJ, Ravasi T, Hume DA. Interferon-gamma: an overview of signals, mechanisms and functions. J Leukoc Biol 2003; 75:163-89. [PMID: 14525967 DOI: 10.1189/jlb.0603252] [Citation(s) in RCA: 3009] [Impact Index Per Article: 136.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Interferon-gamma (IFN-gamma) coordinates a diverse array of cellular programs through transcriptional regulation of immunologically relevant genes. This article reviews the current understanding of IFN-gamma ligand, receptor, signal transduction, and cellular effects with a focus on macrophage responses and to a lesser extent, responses from other cell types that influence macrophage function during infection. The current model for IFN-gamma signal transduction is discussed, as well as signal regulation and factors conferring signal specificity. Cellular effects of IFN-gamma are described, including up-regulation of pathogen recognition, antigen processing and presentation, the antiviral state, inhibition of cellular proliferation and effects on apoptosis, activation of microbicidal effector functions, immunomodulation, and leukocyte trafficking. In addition, integration of signaling and response with other cytokines and pathogen-associated molecular patterns, such as tumor necrosis factor-alpha, interleukin-4, type I IFNs, and lipopolysaccharide are discussed.
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Affiliation(s)
- Kate Schroder
- Institute for Molecular Bioscience, University of Queensland, St. Lucia, Brisbane 4072, Australia.
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50
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Araki T, Tsujioka M, Abe T, Fukuzawa M, Meima M, Schaap P, Morio T, Urushihara H, Katoh M, Maeda M, Tanaka Y, Takeuchi I, Williams JG. A STAT-regulated, stress-induced signalling pathway in Dictyostelium. J Cell Sci 2003; 116:2907-15. [PMID: 12771188 DOI: 10.1242/jcs.00501] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
The Dictyostelium stalk cell inducer differentiation-inducing factor (DIF) directs tyrosine phosphorylation and nuclear accumulation of the STAT (signal transducer and activator of transcription) protein Dd-STATc. We show that hyperosmotic stress, heat shock and oxidative stress also activate Dd-STATc. Hyperosmotic stress is known to elevate intracellular cGMP and cAMP levels, and the membrane-permeant analogue 8-bromo-cGMP rapidly activates Dd-STATc, whereas 8-bromo-cAMP is a much less effective inducer. Surprisingly, however, Dd-STATc remains stress activatable in null mutants for components of the known cGMP-mediated and cAMP-mediated stress-response pathways and in a double mutant affecting both pathways. Also, Dd-STATc null cells are not abnormally sensitive to hyperosmotic stress. Microarray analysis identified two genes, gapA and rtoA, that are induced by hyperosmotic stress. Osmotic stress induction of gapA and rtoA is entirely dependent on Dd-STATc. Neither gene is inducible by DIF but both are rapidly inducible with 8-bromo-cGMP. Again, 8-bromo-cAMP is a much less potent inducer than 8-bromo-cGMP. These data show that Dd-STATc functions as a transcriptional activator in a stress-response pathway and the pharmacological evidence, at least, is consistent with cGMP acting as a second messenger.
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Affiliation(s)
- Tsuyoshi Araki
- School of Life Sciences, University of Dundee, Wellcome Trust Biocentre, Dow Street, Dundee, DD1 5EH, UK
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