关键词： ATF3   Nitric oxide   iNOS   NF-kappa B: macrophages  

摘要： Macrophage-associated nitric oxide (NO) production plays a crucial role in the pathogenesis of tissue damage. However, negative factors that regulate NO production remains poorly understood despite its significance of NO homeostasis. Here, we show that activating transcription factor 3 (ATF3), a transcriptional regulator of cellular stress responses, was strongly induced in activated macrophages and its depletion resulted in pronounced enhancement of inducible nitric oxide synthase (iNOS) gene expression and subsequently the induction of high levels of NO production. In response to lipopolysaccharide (LPS) and IFN-gamma, ATF3 inhibited transcriptional activity of NF-kappa B by interacting with the N-terminal (1-200 amino acids) of p65 and was bound to the NF-kappa B promoter, leading to suppression of iNOS gene expression. In addition, inhibitory effects of ATF3 on iNOS and NO secretion were suppressed by inhibitor of casein kinase II (CK2) activity or its knockdown. Moreover, the levels of ATF3 were highly elevated in established cecal ligation and puncture or LPS-injected mice, a model of endotoxemia. ATF3 is also elevated in peritoneal macrophages. Collectively, our findings suggest that ATF3 regulates NO homeostasis by associating with NF-kappa B component, leading to the repression of its transcriptional activity upon inflammatory signals and points to its potential relevance for the control of cell injuries mediated by NO during macrophage activation.

关键词： Gene regulation   Apoptosis   Akt   NF-kappa B   CREB  

摘要： Activating transcription factor 3 (ATF3) is a stress-induced transcription factor with diverse functions under disease states in multiple cell types. ATF3 has neuroprotective action against cerebral ischemia, which may involve caspase 3. However, the molecular mechanisms underlying ATF3 regulation of apoptosis are largely unknown. Here, we used gain- and loss-of-function and rescue approaches to demonstrate ATF3 attenuating hypoxic neuronal apoptosis. As well, the protective effect of ATF3 was mediated by downregulation of carboxyl-terminal modulator protein (CTMP), a pro-apoptotic factor that inhibits the anti-apoptotic Akt/PKB cascade. ATF3 (1) downregulated the mRNA and protein levels of CTMP;(2) its temporal expression pattern was reciprocal to that of CTMP;and (3) nuclear localization suggested that ATF3 may regulate CTMP transcription following hypoxic insult. Reporter assays demonstrated that ATF3 suppressed CTMP transcription, whereas ATF3 fusion with VP16, converting ATF3 to transcriptional activator, boosted CTMP transcription. By contrast, NF-kappa B increased CTMP transcription, and degradation-resistant I kappa B alpha decreased CTMP transcription. ChIP assays further confirmed that binding of ATF3 to the ATF/CREB site hindered NF-kappa B binding to the CTMP promoter, which repressed CTMP expression. Furthermore, CTMP siRNA treatment reduced hypoxic neuronal apoptosis by increasing p-Akt (Ser473) levels and leaving the upstream ATF3 level unchanged. We have identified an endogenous neuroprotective ATF3 -> CTMP signal cascade that may be a therapeutic target for reducing ischemic brain injury.

关键词： DNA damage and repair   Stress signalling  

摘要： Tat-interactive protein 60 (Tip60) is a MYST histone acetyltransferase that catalyses acetylation of the major DNA damage kinase Ataxia telangiectasia mutated (ATM), thereby triggering cellular signalling required for the maintenance of genomic stability on genotoxic insults. The Tip60 activity is modulated by post-translational modifications that alter its stability and its interactions with substrates. Here we report that activating transcription factor 3 (ATF3), a common stress mediator and a p53 activator, is a regulator of Tip60. ATF3 directly binds Tip60 at a region adjacent to the catalytic domain to promote the protein acetyltransferase activity. Moreover, the ATF3-Tip60 interaction increases the Tip60 stability by promoting USP7-mediated deubiquitination of Tip60. Consequently, knockdown of ATF3 expression leads to decreased Tip60 expression and suppression of ATM signalling as evidenced by accumulated DNA lesions and increased cell sensitivity to irradiation. Our findings thus reveal a previously unknown function of a common stress mediator in regulating Tip60 function.

关键词： ARTIFICIAL respiration -- Complications   LUNGS -- Wounds & injuries -- Treatment   TRANSCRIPTION factors   DISEASE susceptibility   PROTEIN expression   CELLULAR signal transduction  

摘要： Aims: Ventilator-induced lung injury (VILI) contributes to mortality in patients with acute respiratory distress syndrome, the most severe form of acute lung injury (ALI). Absence of activating transcription factor 3 (ATF3) confers susceptibility to ALI/VILI. To identify cell-specific ATF3-dependent mechanisms of susceptibility to ALI/VILI, we generated ATF3 chimera by adoptive bone marrow (BM) transfer and randomized to inhaled saline or lipopolysacharide (LPS) in the presence of mechanical ventilation (MV). Adenovirus vectors to silence or overexpress ATF3 were used in primary human bronchial epithelial cells and murine BM-derived macrophages from wild-type or ATF3-deficient mice. Results: Absence of ATF3 in myeloid-derived cells caused increased pulmonary cellular infiltration. In contrast, absence of ATF3 in parenchymal cells resulted in loss of alveolar-capillary membrane integrity and increased exudative edema. ATF3-deficient macrophages were unable to limit the expression of pro-inflammatory mediators. Knockdown of ATF3 in resident cells resulted in decreased junctional protein expression and increased paracellular leak. ATF3 overexpression abrogated LPS induced membrane permeability. Despite release of ATF3-dependent Nrf2 transcriptional inhibition, mice that lacked ATF3 expression in resident cells had increased Nrf2 protein degradation. Innovation: In our model, in the absence of ATF3 in parenchymal cells increased Nrf2 degradation is the result of increased Keap-1 expression and loss of DJ-1 (Parkinson disease [autosomal recessive, early onset] 7), previously not known to play a role in lung injury. Conclusion: Results suggest that ATF3 confers protection to lung injury by preventing inflammatory cell recruitment and barrier disruption in a cell-specific manner, opening novel opportunities for cell specific therapy for ALI/VILI. Antioxid. Redox Signal. 22, 651-668.

关键词： gluconeogenesis   glucagon   CREB   ATF3   cAMP  

摘要： Increases in circulating glucagon during fasting maintain glucose balance by stimulating hepatic gluconeogenesis. Acute ethanol intoxication promotes fasting hypoglycemia through an increase in hepatic NADH, which inhibits hepatic gluconeogenesis by reducing the conversion of lactate to pyruvate. Here we show that acute ethanol exposure also lowers fasting blood glucose concentrations by inhibiting the CREB-mediated activation of the gluconeogenic program in response to glucagon. Ethanol exposure blocked the recruitment of CREB and its coactivator CRTC2 to gluconeogenic promoters by up-regulating ATF3, a transcriptional repressor that also binds to cAMP-responsive elements and thereby down-regulates gluconeogenic genes. Targeted disruption of ATF3 decreased the effects of ethanol in fasted mice and in cultured hepatocytes. These results illustrate how the induction of transcription factors with overlapping specificity can lead to cross-coupling between stress and hormone-sensitive pathways.

关键词： ATF3   p21   HDAC   Growth inhibition   Epidermoid carcinoma cell  

摘要： Inhibition of histone deacetylase (HDAC) activity by HDAC inhibitors (HDACis) results in cancer cell growth inhibition, and HDACis have been revealed as potential anti-skin cancer agents. p21 is a cyclin-dependent kinase inhibitor and an essential regulator of growth inhibition. Recently, we reported that activating transcription factor 3 (ATF3) could significantly promote skin cancer cell growth. This study explored the relationship between ATF3 and HDACi-induced growth inhibition of epidermoid carcinoma cells. We found that trichostatin A (TSA) treatment inhibited cell growth in A431 epidermoid carcinoma cells in a dose-dependent manner. Simultaneously, p21 and ATF3 expression levels were upregulated and downregulated upon TSA stimulation, respectively. ATF3 overexpression promoted cell growth and downregulated p21 expression. In contrast, ATF3 depletion resulted in cell growth reduction and p21 transcriptional upregulation. More importantly, ATF3 overexpression partially antagonized TSA-induced growth inhibition and p21 activation. Collectively, these data demonstrate that ATF3 acts as an essential negative regulator of TSA-induced cell growth inhibition through interfering with TSA-induced p21 activation.

关键词： Activating transcription factor 3 (ATF3)   Peroxisome proliferator activated receptor (PPAR gamma)   PPAR gamma responsive element (PPRE)   Transcriptional coactivation   p300  

摘要： Previously, we reported that activating transcription factor 3 (ATF3) downregulates peroxisome proliferator activated receptor (PPAR gamma) gene expression and inhibits adipocyte differentiation in 3T3-L1 cells. Here, we investigated another role of ATF3 on the regulation of PPAR gamma activity. ATF3 inhibited PPAR gamma-stimulated transactivation of PPAR gamma responsive element (PPRE)-containing reporter or GAL4/PPAR gamma chimeric reporter. Thus, ATF3 effectively repressed rosiglitazone-stimulated expression of adipocyte fatty acid binding protein (aP2), PPAR gamma target gene, in 3T3-L1 cells. Coimmunoprecipitation and GST pulldown assay demonstrated that ATF3 interacted with PPAR gamma. Accordingly, ATF3 prevented PPAR gamma from binding to PPRE on the aP2 promoter. Furthermore, ATF3 suppressed p300-mediated transcriptional coactivation of PPRE-containing reporter. Chromatin immunoprecipitation assay showed that overexpression of ATF3 blocked both binding of PPAR gamma and recruitment of p300 to PPRE on aP2 promoter induced by rosiglitazone treatment in 3T3-L1 cells. Taken together, these results suggest that ATF3 interacts with PPAR gamma and represses PPAR gamma-mediated transactivation through suppression of p300-stimulated coactivation in 3T3-L1 cells, which may play a role in inhibition of adipocyte differentiation. (C) 2014 Elsevier Inc. All rights reserved.

关键词： spermatocytes   Ahr   Nrf2   CSC   ATF3   E2F4   p38 MAPK   and ERK  

摘要： Our earlier studies have demonstrated that the cigarette smoke in the form of cigarette smoke condensate(CSC)causes growth arrest of a mouse spermatocyte cell line[GC-2spd(ts)]through activation of the AHR–NRF2 *** present study demonstrates the CSC-activated p38 and ERK MAPK signaling in GC-2spd(ts)via arylhydrocarbon receptor(AHR).Pharmacological inhibition by using AHR-antagonist,or p38 MAPK and ERK(MEK1)inhibitors significantly abrogates CSC-induced growth arrest by AHR and MAPK ***-PCR,western blot,and immunofluorescence of Ahr-target of Nrf2,and stress-inducible growth suppressive Atf3 and E2f4 following treatments indicate a crosstalk among these *** of Atf3 by Nrf2 and Ahr through RNA interference suggests the existence of a cross-regulatory loop between the *** induction of E2f4 via Atf3 and its regulation by pharmacological inhibitors reveal a possible regulatory mechanism of growth inhibitory *** silencing of Ahr,Nrf2,Atf3,and E2f4 genes and downregulation of cyclins by CSC corroborate the growth inhibitory effect of cigarette ***,the data obtained suggest that the CSC-mediated MAPKs and AHR–NRF2 crosstalks lay the molecular basis for the growth arrest and cell death of spermatocytes.

关键词： 活化转录因子3   脊髓损伤   斑马鱼模型   炎症因子   轴突再生   神经发育  

摘要： 背景：脊髓损伤是一种严重的中枢神经系统致残性疾病,目前针对脊髓损伤尚无有效的治疗手段和药物。哺乳动物的神经系统拥有非常有限的修复和再生能力。与哺乳动物不同，斑马鱼在脊髓损伤后具有很强的修复和再生能力，成年斑马鱼在脊髓损伤后六周时间内可基本恢复正常运动水平，但其具体机制尚不明确。已有研究证明活化转录因子3（ATF3）在斑马鱼视神经损伤后可以促进视神经细胞再生和修复。这提示我们，在斑马鱼脊髓损伤后，ATF3分子可能对神经再生和修复起到重要作用。\n 目的：本课题利用成年斑马鱼脊髓横断模型，探究ATF3在脊髓损伤后的mRNA和蛋白水平的变化。同时在抑制ATF3蛋白表达后，研究其对斑马鱼脊髓修复的影响和相关机制。\n 方法：选用4-6月龄的成年斑马鱼进行脊髓横断模型手术，在损伤后不同时间点取损伤点下段4mm的脊髓组织进行实时定量PCR，原位杂交以及免疫荧光，检测ATF3 mRNA和蛋白变化，同时在脊髓损伤后不同时间点用免疫荧光共标ATF3与运动神经元或ATF3与放射状胶质细胞。斑马鱼脊髓损伤后，用ATF3 morpholino（MO）抑制斑马鱼脊髓损伤后ATF3蛋白质表达，检测斑马鱼运动恢复情况以及穿过损伤位点神经轴突修复的水平变化，同时用实时定量 PCR的方法检测肿瘤坏死因子-α（TNF-α）和白介素-1β（IL-1β）mRNA的表达水平变化以探究炎症因子对修复的影响。最后，利用显微注射技术将ATF3 MO注射到斑马鱼胚胎中，探究ATF3分子对斑马鱼幼鱼发育的影响。\n 结果：\n ***3 mRNA水平在斑马鱼脊髓损伤后的4小时、12小时和6天维持在较高的表达水平，而在损伤后11天，其恢复到假手术组的表达水平。\n ***3蛋白水平在脊髓损伤后的4小时、12小时和6天表达水平升高，11天降低到假手术组水平。\n ***3蛋白表达在运动神经元的细胞浆中，并且在脊髓损伤后12小时和6天表达水平增高。\n ***3蛋白毗邻但不表达在放射状胶质细胞中，在脊髓损伤后6天，放射状胶质细胞数量增加。\n ***3 MO抑制ATF3蛋白在斑马鱼脊髓内表达后，脊髓损伤后第5和第6周，抑制组斑马鱼运动能力分别降低了36.4%和35.7%。\n 6.通过神经示踪技术发现，抑制脊髓ATF3表达六周后，神经轴突修复被抑制。\n 7.抑制脊髓ATF3表达11天后，相比MO对照组，抑制组TNF-α和IL-1βmRNA表达水平维持在高水平，在脊髓损伤后21天，恢复到对照组水平。\n ***3 MO抑制ATF3蛋白在斑马鱼胚胎发育中的表达后，幼鱼发育畸形，并且在受精5天后出现死亡率升高和身体长度变短等现象。\n 结论：ATF3分子通过调控炎症因子和轴突再生参与斑马鱼脊髓损伤后的修复，并对幼鱼的正常发育起到重要作用。

关键词： ACRYLAMIDE   MACROPHAGES   CELLULAR aging   ACTIVE oxygen -- Physiological effect   TELOMERASE   JNK mitogen-activated protein kinases   CELL cycle -- Regulation  

摘要： Senescence, which is irreversible cell cycle arrest, is induced by various types of DNA damage, including genotoxic stress. Senescent cells show dysregulation of tumor suppressor genes and other regulators of cellular proliferation. Activating transcription factor 3 (ATF3) plays a pleiotropic role in biological processes through genotoxic stress. In this study, we examined the effects of acrylamide (ACR), a genotoxic carcinogen, on cellular senescence and the molecular mechanisms of ATF3 function in macrophages. Treatment of macrophages with ACR at low concentrations (<1.0 mM) resulted in senescence-like morphology and an increase in senescence-associated beta-galactosidase (SA-beta-gal) activity. Exposure of macrophages to ACR led to stress-induced, telomerase-independent senescence. In addition, ACR treatment for 1, 3, or 5 days showed a concentration-dependent increase in ATF3 expression and G0/G1 phase arrest. To better understand the role of ATF3 in controlling the senescence response to ACR, SA-beta-gal activity was examined using ATF3 knockdown and overexpression. ACR-mediated senescence was significantly decreased by knockdown of ATF3, whereas it was increased with ATF3 overexpression. We found that ATF3 regulated p53 and p21 levels. ATF3 also played an important role in regulating intracellular reactive oxygen species (ROS) production in response to ACR treatment. Moreover, phosphorylation of p38 and JNK kinases, which were activated during ATF3-mediated senescence, was observed in ACR-treated macrophages. Taken together, these results suggest that ATF3 contributes to ACR-induced senescence by enhancing ROS production, activating p38 and JNK kinases, and promoting the ATF3-dependent expression of p53, resulting in regulation of cellular senescence in macrophages.