Activating Transcription Factor 3 Gene Research Articles

The effect of feeding with different protein levels on internal organ weight and gene expression of MEF2A and ATF3 in crossbred local chicken using RT-PCR

BackgroundMyogenic enhancer transcription factor 2A (MEF2A) is a transcription factor known for its role in controlling skeletal muscle regeneration and metabolic processes, while activating transcription factor 3 (ATF3) is a stress-induced transcription factor that plays a role in modulating metabolic processes, immunity, and oncogenesis. Environmental factors, such as dietary protein, can influence gene expression levels. Insufficient protein intake can negatively affect the metabolic performance of internal organs, leading to the abnormal weight of internal organs. A total of 192 non-sexing crossbred local chickens day-old-chick (DOC) with a completely randomized design (CRD) method of 3 treatments and 8 replicates. Real-time polymerase chain reaction (RT-PCR) is used to measure the gene expression levels. This study aimed to determine the effect of feeding with various protein levels on internal organ weight and gene expression of MEF2A and ATF3 in crossbred local chickens. ResultThe analysis of treatment revealed that the results were not significantly different (P > 0.05) on gizzard weight and spleen weight. However, it shows a significantly different result (P < 0.05) on heart weight and a highly significantly different result (P < 0.01) on pancreas weight. These findings suggest that protein levels in the diet had a significant impact on heart and pancreas weights. In terms of gene expression, the increased utilization of protein did not result in an elevation of MEF2A gene expression in both muscle tissue and liver tissue. Specifically, in muscle tissue, MEF2A gene expression was highly expressed at 18% protein feed for the starter phase and 16% for the finisher phase. Conversely, in liver tissue, MEF2A gene expression was highly expressed at 22% protein feed for the starter phase and 20% for the finisher phase. Moreover, ATF3 gene expression in muscle tissue exhibited a negative correlation with increasing feed protein levels. ConclusionThe results indicate that varying protein levels did not lead to abnormal weights in the liver, kidney, heart, and spleen organs. Additionally, the differential gene expression patterns of MEF2A and ATF3 in muscle tissue and liver tissue suggest that these genes respond differently to varying protein-feeding treatments. These findings provide insights into the complex regulatory mechanisms of MEF2A and ATF3 genes in relation to protein levels and organ-specific responses in crossbred local chickens.

Decreased expression of ATF-3 in melanoma promotes tumor growth via ERK and AKT pathways

<h3>Introduction</h3> Activating transcription factor 3 (ATF-3), a cyclic AMP-dependent transcription factor that is encoded by the ATF-3 gene, plays a regulatory role in melanoma, although its function during tumor progression is unknown. Here we evaluated ATF-3 expression during melanoma evolution to elucidate its role in melanogenesis. <h3>Materials and Methods</h3> Primary and metastatic melanoma cell lines, patient melanomas and murine melanoma xenografts were evaluated by immunofluorescence, Western Blot and RT-PCR studies. Melanoma cell lines were retrovirally transfected with ATF-3 overexpressing vectors and CCK8, wound healing and Matrigel invasion studies were performed to test for proliferation, migration and invasive properties, respectively. Xenograft murine models were designed by subcutaneously injecting melanoma cells or its overexpressed counterpart into the anterior and posterior flanks of nude mice to evaluate tumor size and rate of formation. <h3>Results</h3> By immunofluorescence, ATF-3 staining declined in metastatic versus primary melanoma cell lines and with progression from patient nevi to primary to metastatic melanomas. Analysis of TCGA skin cutaneous melanoma database showed that lower ATF3 expression correlated with poor prognosis. Metastatic melanoma cell lines with retrovirally-induced ATF-3 overexpression (A2058 and C8161 ATF-3 OE) exhibited decreased prolif- eration, migration and invasion. GO and KEGG analyses of our RNA seq data showed downregulation of phospho- rylated ERK and AKT in A2058 ATF-3 OE cells that was further validated by Western Blot. In vivo, xenografted A2058 ATF-3 OE cells formed smaller and less abundant tumors in murine subcutis than did control cells, and Ki-67 staining confirmed lower labeling indices in ATF-3 overexpressing lesions. <h3>Conclusions</h3> ATF-3 expression is associated with melanoma virulence and thus provides a potential target for novel therapies and predictive biomarker applications.

Hepatic IRF2BP2 Mitigates Nonalcoholic Fatty Liver Disease by Directly Repressing the Transcription of ATF3

Although knowledge regarding the pathogenesis of nonalcoholic fatty liver disease (NAFLD) has profoundly grown in recent decades, the internal restrictive mechanisms remain largely unknown. We have recently reported that the transcription repressor interferon regulatory factor-2 binding protein 2 (IRF2BP2) is enriched in cardiomyocytes and inhibits pathological cardiac hypertrophy in mice. Notably, IRF2BP2 is abundantly expressed in hepatocytes and dramatically down-regulated in steatotic livers, whereas the role of IRF2BP2 in NAFLD is unknown. Herein, using gain-of-function and loss-of-function approaches in mice, we demonstrated that while hepatocyte-specific Irf2bp2 knockout exacerbated high-fat diet-induced hepatic steatosis, insulin resistance and inflammation, hepatic Irf2bp2 overexpression protected mice from these metabolic disorders. Moreover, the inhibitory role of IRF2BP2 on hepatosteatosis is conserved in a human hepatic cell line in vitro. Combinational analysis of digital gene expression and chromatin immunoprecipitation sequencing identified activating transcription factor 3 (ATF3) to be negatively regulated by IRF2BP2 in NAFLD. Chromatin immunoprecipitation and luciferase assay substantiated the fact that IRF2BP2 is a bona fide transcription repressor of ATF3 gene expression via binding to its promoter region. Functional studies revealed that ATF3 knockdown significantly relieved IRF2BP2 knockout-exaggerated hepatosteatosis in vitro. IRF2BP2 is an integrative restrainer in controlling hepatic steatosis, insulin resistance, and inflammation in NAFLD through transcriptionally repressing ATF3 gene expression.

Transcription factor ATF3 mediates the radioresistance of breast cancer.

This study was designed to research the influence of activating transcription factor 3 (ATF3) on the radioresistance of breast cancer. ATF3 expression was measured by qRT‐PCR and immunohistochemistry. Cancerous cell lines were cultured in vitro, and the expression of ATF3 was gauged by both qRT‐PCR and Western blot before and after the radiation therapy. Cellular cycle and apoptosis were analysed by flow cytometry. Changes in the expression of corresponding proteins in the downstream pathways were identified by Western blot. Tumour xenograft was used to evaluate the effect of ATF3 in vivo. ATF3 was observed stronger in breast cancer tissues and cells. After radiation therapy, the expression of ATF3 in breast cancer cells was up‐regulated. Silencing ATF3 could promote G2/M phase block, facilitate cell apoptosis and decrease clonogenic survival rate. The overexpression of ATF3 could curb G2/M‐phase block, cell apoptosis and increase clonogenic survival rate. Overexpression ATF3 could increase radioresistance by up‐regulating the level of phosphorylation of Akt in the PI3K/Akt signalling pathway. Radioresistance of breast cancer cells could be alleviated by inhibiting the PI3K/Akt signalling pathway. ATF3 could also promote radioresistance in vivo. ATF3 gene was able to promote radioresistance of breast cancer cells.

Carnosol suppresses interleukin-6 production in mouse lungs injured by ischemia-reperfusion operation and in RAW264.7 macrophages treated with lipopolysaccharide.

Carnosol is a naturally occurring herbal compound, known for its antioxidative properties. We previously found that carnosol protected mouse lungs from ischemia-reperfusion injury in ex vivo cultures. To elucidate the molecular mechanisms underpinning carnosol-mediated lung protection, we analyzed modes of interleukin-6 (IL-6) gene expression, which is associated with lung ischemia-reperfusion injury. Microarray analysis of mouse lungs suggested that IL-6 mRNA levels were elevated in the mouse lungs subjected to clamp-reperfusion, which was associated with elevated levels of other inflammatory modulators, such as activating transcription factor 3 (ATF3). Carnosol pretreatment lowered the IL-6 protein levels in mouse lung homogenates prepared after the clamp-reperfusion. On the other hand, the ATF3 gene expression was negatively correlated with that of IL-6 in RAW264.7 cells. IL-6 mRNA levels and gene promoter activities were suppressed by carnosol in RAW264.7 cells, but rescued by ATF3 knockdown. When RAW264.7 cells were subjected to hypoxia-reoxygenation, carnosol treatment lowered oxygen consumption after reoxygenation, which was coupled with a correlation with a transient production of mitochondrial reactive oxygen species and following ATF3 gene expression. These results suggest that carnosol treatment could be a new strategy for protecting lungs from ischemia-reperfusion injury by modulating the ATF3-IL-6 axis.

Larval hemolymph of rhinoceros beetle, Allomyrina dichotoma, enhances insulin secretion through ATF3 gene expression in INS-1 pancreatic β-cells.

Abstract In this study, we show that INS-1 pancreatic β-cells treated for 2 h with hemolymph of larvae of rhinoceros beetle, Allomyrina dichotoma, secreted about twice as much insulin compared to control cells without such treatment. Activating transcription factor 3 (ATF3) was the highest upregulated gene in DNA chip analysis. The A. dichotoma hemolymph dose-dependently induced increased expression levels of genes encoding ATF3 and insulin. Conversely, treatment with ATF3 siRNA inhibited expression levels of both genes and curbed insulin secretion. These results suggest that the A. dichotoma hemolymph has potential for treating and preventing diabetes or diabetes-related complications.

Activating transcription factor 3 promotes intestinal epithelial cell apoptosis in Crohn’s disease

Intestinal epithelial cell (IEC) apoptosis plays a vital role in the pathogenesis of Crohn’s disease (CD), which is an inflammatory bowel disease (IBD). Activating transcription factor 3 (ATF3) modulates apoptosis under stress via regulating the p53 pathway. However, the expression and function of ATF3 in CD are unclear. In the present study, ATF3, p53, and p53 target gene Bax expression increased in CD patients; a mouse 2, 4, 6-trinitrobenzenesulfonic acid (TNBS)-induced CD model; and a TNF-α-treated HT29 cell colitis model. ATF3 knockdown effectively decreased TNF-α-induced p53 and Bax expression, as well as inhibited the apoptosis of HT29 cells. Additionally, ATF3 enhanced the stability and transcription activity of p53 via interacting with p53. In summary, these data indicated that ATF3 might promote IEC apoptosis in CD via up-regulating the stability and transcription activity of p53, implying a novel molecular target for CD therapy.

Regulation of the ATF3 gene by a single promoter in response to amino acid availability and endoplasmic reticulum stress in human primary hepatocytes and hepatoma cells

Activating transcription factor 3 (ATF3) is a highly regulated protein that is implicated in a wide range of pathological conditions including inflammation and transformation. Transcription from the ATF3 gene is induced by several stress-induced signaling pathways, including amino acid limitation (amino acid response, AAR) and ER stress (unfolded protein response, UPR). Induction of ATF3 transcription by these pathways is mediated by ATF4 and cJUN recruitment to enhancer elements within the ATF3 gene. Although a canonical promoter (promoter A) has been studied by numerous laboratories, a second promoter activity (promoter A1), 43 kb upstream of the first, has been reported to respond to stress-induced signaling and to be critical for ATF3 expression in certain transformed cells. The results of the present study show that in normal human hepatocytes and HepG2 human hepatoma cells both basal as well as AAR- and UPR-induced transcription occurs almost exclusively from promoter A. This selectivity between the two promoters correlated with increased binding of ATF4, recruitment of RNA polymerase II, and the expected histone modifications in the promoter A region of the gene. Time course studies of ATF3 transcription activity revealed that the temporal kinetics for ATF3 induction differ between the AAR and UPR, with the former being more transient than the latter. Collectively, the results document that ATF3 expression in normal and transformed human liver originates from the canonical promoter A that responds to multiple stress signals.

ATF3-Dependent Regulation of EGR1 in vitro and in vivo

Background/Aims: Activating transcription factor 3 (ATF3) and early growth response protein 1 (EGR1) are reported to interact, but their use as prognostic factors in cancer is discussed controversially. Methods: We measured ATF3 and EGR1 gene expression changes in human mini-organ cultures (MOCs) of healthy nasal epithelia, UM-SCC-22B, and FADU_DD cells after acid reflux exposure. Next, ATF3 and EGR1 gene expression was analysed in tumour tissues and related to the median expression of autologous reference tissue samples. Results: ATF3 and EGR1 mRNA expression was significantly reduced after consecutive exposure of MOCs at pH <7.0 to artificial gastric juice (refluxate). In contrast, ATF3 mRNA was upregulated significantly within the first hour of incubation. EGR1 mRNA exhibited no significant changes. The analysed cell lines exhibited a cell line-specific alteration. In FADU_DD cells, the upregulation of EGR1 was significant after refluxate exposure, but in HN-SCC 22B, no significant changes were detected. The analysis of the HNSCC samples confirmed the heterogeneous data of the literature. Conclusion: The data maintain the hypothesis that ATF3 and EGR1 are involved in the beginning of inflammatory processes. Whether these two transcription factors act as tumour suppressors or promoters is context dependent and warrants analysis in further studies.

Induction of Activating Transcription Factor 3 Is Associated with Cisplatin Responsiveness in Non–Small Cell Lung Carcinoma Cells

Non–small cell lung carcinoma (NSCLC) is the most common cause of cancer deaths, with platin-based combination chemotherapy the most efficacious therapies. Gains in overall survival are modest, highlighting the need for novel therapeutic approaches including the development of next-generation platin combination regimens. The goal of this study was to identify novel regulators of platin-induced cytotoxicity as potential therapeutic targets to further enhance platin cytotoxicity. Employing RNA-seq transcriptome analysis comparing two parental NSCLC cell lines Calu6 and H23 to their cisplatin-resistant sublines, Calu6cisR1 and H23cisR1, activating transcription factor 3 (ATF3) was robustly induced in cisplatin-treated parental sensitive cell lines but not their resistant sublines, and in three of six tumors evaluated, but not in their corresponding normal adjacent lung tissue (0/6). Cisplatin-induced JNK activation was a key regulator of this ATF3 induction. Interestingly, in both resistant sublines, this JNK induction was abrogated, and the expression of an activated JNK construct in these cells enhanced both cisplatin-induced cytotoxicity and ATF3 induction. An FDA-approved drug compound screen was employed to identify enhancers of cisplatin cytotoxicity that were dependent on ATF3 gene expression. Vorinostat, a histone deacetylase inhibitor, was identified in this screen and demonstrated synergistic cytotoxicity with cisplatin in both the parental Calu6 and H23 cell lines and importantly in their resistant sublines as well that was dependent on ATF3 expression. Thus, we have identified ATF3 as an important regulator of cisplatin cytotoxicity and that ATF3 inducers in combination with platins are a potential novel therapeutic approach for NSCLC.

Lipotoxic brain microvascular injury is mediated by activating transcription factor 3-dependent inflammatory and oxidative stress pathways

Dysfunction of the cerebrovasculature plays an important role in vascular cognitive impairment (VCI). Lipotoxic injury of the systemic endothelium in response to hydrolyzed triglyceride-rich lipoproteins (TGRLs; TGRL lipolysis products) or a high-fat Western diet (WD) suggests similar mechanisms may be present in brain microvascular endothelium. We investigated the hypothesis that TGRL lipolysis products cause lipotoxic injury to brain microvascular endothelium by generating increased mitochondrial superoxide radical generation, upregulation of activating transcription factor 3 (ATF3)-dependent inflammatory pathways, and activation of cellular oxidative stress and apoptotic pathways. Human brain microvascular endothelial cells were treated with human TGRL lipolysis products that induced intracellular lipid droplet formation, mitochondrial superoxide generation, ATF3-dependent transcription of proinflammatory, stress response, and oxidative stress genes, as well as activation of proapoptotic cascades. Male apoE knockout mice were fed a high-fat/high-cholesterol WD for 2 months, and brain microvessels were isolated by laser capture microdissection. ATF3 gene transcription was elevated 8-fold in the hippocampus and cerebellar brain region of the WD-fed animals compared with chow-fed control animals. The microvascular injury phenotypes observed in vitro and in vivo were similar. ATF3 plays an important role in mediating brain microvascular responses to acute and chronic lipotoxic injury and may be an important preventative and therapeutic target for endothelial dysfunction in VCI.

The catalytic topoisomerase II inhibitor dexrazoxane induces DNA breaks, ATF3 and the DNA damage response in cancer cells.

The catalytic topoisomerase II inhibitor dexrazoxane has been associated not only with improved cancer patient survival but also with secondary malignancies and reduced tumour response. We investigated the DNA damage response and the role of the activating transcription factor 3 (ATF3) accumulation in tumour cells exposed to dexrazoxane. Dexrazoxane exposure induced topoisomerase IIα (TOP2A)-dependent cell death, γ-H2AX accumulation and increased tail moment in neutral comet assays. Dexrazoxane induced DNA damage responses, shown by enhanced levels of γ-H2AX/53BP1 foci, ATM (ataxia telangiectasia mutated), ATR (ATM and Rad3-related), Chk1 and Chk2 phosphorylation, and by p53 accumulation. Dexrazoxane-induced γ-H2AX accumulation was dependent on ATM. ATF3 protein was induced by dexrazoxane in a concentration- and time-dependent manner, which was abolished in TOP2A-depleted cells and in cells pre-incubated with ATM inhibitor. Knockdown of ATF3 gene expression by siRNA triggered apoptosis in control cells and diminished the p53 protein level in both control and dexrazoxane -treated cells. This was accompanied by increased γ-H2AX accumulation. ATF3 knockdown also delayed the repair of dexrazoxane -induced DNA double-strand breaks. As with other TOP2A poisons, dexrazoxane induced DNA double-strand breaks followed by activation of the DNA damage response. The DNA damage-triggered ATF3 controlled p53 accumulation and generation of double-strand breaks and is proposed to serve as a switch between DNA damage and cell death following dexrazoxane treatment. These findings suggest a mechanistic explanation for the diverse clinical observations associated with dexrazoxane.

Inhibition of kinase and endoribonuclease activity of ERN1/IRE1α affects expression of proliferationrelated genes in U87 glioma cells

AbstractInhibition of ERN1/IRE1α (endoplasmic reticulum to nucleus signaling 1/inositol requiring enzyme-1α), the major signaling pathway of endoplasmic reticulum stress, significantly decreases tumor growth. We have studied the expression of transcription factors such as E2F8 (E2F transcription factor 8), EPAS1 (endothelial PAS domain protein 1), TBX3 (T-box 3), ATF3 (activating transcription factor 3), FOXF1 (forkhead box F1), and HOXC6 (homeobox C6) in U87 glioma cells overexpressing dominant-negative ERN1/IRE1α defective in endoribonuclease (dnr-ERN1) as well as defective in both kinase and endonuclease (dn-ERN1) activity of ERN1/IRE1α. We have demonstrated that the expression of all studied genes is decreased at the mRNA level in cells with modified ERN1/IRE1α; TBX3, however, is increased in these cells as compared to control glioma cells. Changes in protein levels of E2F8, HOXC6, ATF3, and TBX3 corresponded to changes in mRNAs levels. We also found that two mutated ERN1/IRE1α have differential effects on the expression of studied transcripts. The presence of kinase and endonuclease deficient ERN1/IRE1α in glioma cells had a less profound effect on the expression of E2F8, HOXC6, and TBX3 genes than the blockade of the endoribonuclease activity of ERN1/IRE1α alone. Kinase and endonuclease deficient ERN1/IRE1α suppresses ATF3 and FOXF1 gene expressions, while inhibition of only endoribonuclease of ERN1/IRE1α leads to the up-regulation of these gene transcripts. The present study demonstrates that fine-tuning of the expression of proliferation related genes is regulated by ERN1/IRE1α an effector of endoplasmic reticulum stress. Inhibition of ERN1/IRE1α, especially its endoribonuclease activity, correlates with deregulation of proliferation related genes and thus slower tumor growth.

Temporal changes in neurotrophic factors and neurite outgrowth in the major pelvic ganglion following cavernous nerve injury.

Despite nerve-sparing radical prostatectomy, nerve damage and erectile dysfunction (ED) prevail, and preventing neurodegeneration is of great importance. Neurotrophic factors and neurite outgrowth were characterized in major pelvic ganglia (MPG) following bilateral cavernous nerve injury (BCNI). Young male Sprague-Dawley rats underwent sham or BCNI surgery, and the intracavernosal pressure to mean arterial pressure ratio was measured 2, 7, 14, 21, 30, and 60 days following injury (n = 8/group). MPG gene expression (qPCR) and Western blot were performed for glial cell line-derived neurotrophic factor (GDNF), nerve growth factor (NGF), neurturin, neurotrophin (NT)-3, NT4, brain-derived neurotrophic factor (BDNF), vascular endothelial growth factor, and activating transcription factor 3 (ATF3). Additional rats were injured, and MPGs were removed 24 hr, 48 hr, 3 days, and 7 days following BCNI (n = 3/group). MPGs were cultured in Matrigel, and neurite outgrowth was measured. Erections were impaired early and improved by 60 days in BCNI rats. GDNF, NGF, BDNF, and ATF3 gene expression was significantly increased and NT3 was decreased in MPGs following BCNI (48 hr to 21 days, P < 0.05). GDNF and NGF protein levels were elevated in 48-hr BCNI rats. MPG neurite outgrowth from 24-hr and 48-hr BCNI was higher than sham (658 ± 19 μm, 607 ± 24 μm, 393 ± 23 μm, respectively, P < 0.05). Further studies examining the roles of neurotrophic factors in modulating signaling pathways may provide therapeutic avenues for neurogenically mediated ED.

The induction of activating transcription factor 3 (ATF3) contributes to anti-cancer activity of Abeliophyllum distichum Nakai in human colorectal cancer cells.

BackgroundRecently, Abeliophyllum distichum Nakai (A. distichum) has been reported to exert the inhibitory effect on angiotensin converting enzyme. However, no specific pharmacological effects from A. distichum have been described. We performed in vitro study to evaluate anti-cancer properties of A. distichum and then elucidate the potential mechanisms.MethodsCell viability was measured by MTT assay. ATF3 expression level was evaluated by Western blot or RT-PCR and ATF3 transcriptional activity was determined using a dual-luciferase assay kit after the transfection of ATF3 promoter constructs. In addition, ATF3-dependent apoptosis was evaluated by Western blot after ATF3 knockdown using ATF3 siRNA.ResultsExposure of ethyl acetate fraction from the parts of A. distichum including flower, leaf and branch to human colorectal cancer cells, breast cancer cells and hepatocellular carcinoma reduced the cell viability. The branch extracts from A. distichum (EAFAD-B) increased the expression of activating transcription factor 3 (ATF3) and promoter activity, indicating transcriptional activation of ATF3 gene by EAFAD-B. In addition, our data showed that EAFAD-B-responsible sites might be between -147 and -85 region of the ATF3 promoter. EAFAD-B-induced ATF3 promoter activity was significantly decreased when the CREB site was deleted. However, the deletion of Ftz sites did not affect ATF3 promoter activity by EAFAD-B. We also observed that inhibition of p38MAPK and GSK3β attenuated EAFAD-B-mediated ATF3 promoter activation. Also, EAFAD-B contributes at least in part to increase of ATF3 accumulation.ConclusionThese findings suggest that the anti-cancer activity of EAFAD-B may be a result of ATF3 promoter activation and subsequent increase of ATF3 expression.

Anti-cancer activity of Ginger (Zingiber officinale) leaf through the expression of activating transcription factor 3 in human colorectal cancer cells.

BackgroundGinger leaf (GL) has long been used as a vegetable, tea and herbal medicine. However, its pharmacological properties are still poorly understood. Thus, we performed in vitro studies to evaluate anti-cancer properties of ginger leaf and then elucidate the potential mechanisms involved.MethodsCell viability was measured by MTT assay. ATF3 expression level was evaluated by Western blot or RT-PCR and ATF3 transcriptional activity was determined using a dual-luciferase assay kit after the transfection of ATF3 promoter constructs. In addition, ATF3-dependent apoptosis was evaluated by Western blot after ATF3 knockdown using ATF3 siRNA.ResultsExposure of GL to human colorectal cancer cells (HCT116, SW480 and LoVo cells) reduced the cell viability and induced apoptosis in a dose-dependent manner. In addition, GL reduced cell viability in MCF-7, MDA-MB-231 and HepG-2 cells. ATF3 knockdown attenuated GL-mediated apoptosis. GL increased activating transcription factor 3 (ATF3) expressions in both protein and mRNA level and activated ATF3 promoter activity, indicating transcriptional activation of ATF3 gene by GL. In addition, our data showed that GL-responsible sites might be between -318 and -85 region of the ATF3 promoter. We also observed that ERK1/2 inhibition by PD98059 attenuated GL-mediated ATF3 expression but not p38 inhibition by SB203580, indicating ERK1/2 pathway implicated in GL-induced ATF3 activation.ConclusionsThese findings suggest that the reduction of cell viability and apoptosis by GL may be a result of ATF3 promoter activation and subsequent increase of ATF3 expression through ERK1/2 activation in human colorectal cancer cells.

Characterization of the regulatory mechanisms of activating transcription factor 3 by hypertrophic stimuli in rat cardiomyocytes.

AimsActivating transcription factor 3 (ATF3) is a stress-activated immediate early gene suggested to have both detrimental and cardioprotective role in the heart. Here we studied the mechanisms of ATF3 activation by hypertrophic stimuli and ATF3 downstream targets in rat cardiomyocytes.Methods and ResultsWhen neonatal rat cardiomyocytes were exposed to endothelin-1 (ET-1, 100 nM) and mechanical stretching in vitro, maximal increase in ATF3 expression occurred at 1 hour. Inhibition of extracellular signal-regulated kinase (ERK) by PD98059 decreased ET-1– and stretch–induced increase of ATF3 protein but not ATF3 mRNA levels, whereas protein kinase A (PKA) inhibitor H89 attenuated both ATF3 mRNA transcription and protein expression in response to ET-1 and stretch. To characterize further the regulatory mechanisms upstream of ATF3, p38 mitogen-activated protein kinase (MAPK) signaling was investigated using a gain-of-function approach. Adenoviral overexpression of p38α, but not p38β, increased ATF3 mRNA and protein levels as well as DNA binding activity. To investigate the role of ATF3 in hypertrophic process, we overexpressed ATF3 by adenovirus-mediated gene transfer. In vitro, ATF3 gene delivery attenuated the mRNA transcription of interleukin-6 (IL-6) and plasminogen activator inhibitor-1 (PAI-1), and enhanced nuclear factor-κB (NF-κB) and Nkx-2.5 DNA binding activities. Reduced PAI-1 expression was also detected in vivo in adult rat heart by direct intramyocardial adenovirus-mediated ATF3 gene delivery.ConclusionsThese data demonstrate that ATF3 activation by ET-1 and mechanical stretch is partly mediated through ERK and cAMP-PKA pathways, whereas p38 MAPK pathway is involved in ATF3 activation exclusively through p38α isoform. ATF3 activation caused induction of modulators of the inflammatory response NF-κB and Nkx-2.5, as well as attenuation of pro-fibrotic and pro-inflammatory proteins IL-6 and PAI-1, suggesting cardioprotective role for ATF3 in the heart.

Age-Related Brain Expression and Regulation of the Chemokine CCL4/MIP-1β in APP/PS1 Double-Transgenic Mice

The detrimental effect of activation of the chemokine CCL4/MIP-1β on neuronal integrity in patients with HIV-associated dementia has directed attention to the potential role of CCL4 expression and regulation in Alzheimer disease. Here, we show that CCL4 mRNA and protein are overexpressed in the brains of APPswe/PS1ΔE9 (APP/PS1) double-transgenic mice, a model of cerebral amyloid deposition; expression was minimal in brains from nontransgenic littermates or single-mutant controls. Increased levels of CCL4 mRNA and protein directly correlated with the age-related progression of cerebral amyloid-β (Aβ) levels in APP/PS1 mice. We also found significantly increased expression of activating transcription factor 3 (ATF3), which was positively correlated with age-related Aβ deposition and CCL4 in the brains of APP/PS1 mice. Results from chromatin immunoprecipitation-quantitative polymerase chain reaction confirmed that ATF3 binds to the promoter region of the CCL4 gene, consistent with a potential role in regulating CCL4 transcription. Finally, elevated ATF3 mRNA expression in APP/PS1 brains was associated with hypomethylation of the ATF3 gene promoter region. These observations prompt the testable hypothesis for future study that CCL4 overexpression, regulated in part by hypomethylation of the ATF3 gene, may contribute to neuropathologic progression associated with amyloid deposition in Alzheimer disease.

Genistein‐induced histone modifications on ATF3 may contribute to cell cycle arrest and apoptosis in human colon cancer cell line SW620 (800.8)

Genistein, a soy‐derived isoflavone, has been shown to suppress colon cancer progression by regulating different cellular properties. The aim of the study is to investigate the role of Activating Transcription Factor 3 (ATF3), an immediate‐early response gene, in genistein‐modulated cell cycle arrest and apoptosis in a human colon cancer cell line SW620. Results showed that genistein induced G2 phase arrest and increased late apoptosis in SW620. Genistein also increased the expression of ATF3 mRNA and protein, as well as its transcriptional activity in SW620. mRNA expression of downstream genes of ATF3, such as cell cycle inhibitors p53, p21, CyclinD1, and apoptotic enzymes Casp8, and Casp9, were all induced by genistein. Chromatin immunoprecipitation (ChIP) showed decreased phosphorylated histone H3 serine 10 (H3S10p) at both the promoter and coding region of ATF3. In addition, genistein increased trimethylated histone H3 lysine 36 (H3K36me3) level at a coding region and dimethylated histone H3 lysine 4 (H3K4me2) level at the promoter region. Methylation sensitive PCR showed that genistein did not change the level of DNA methylation at various regions of ATF3 gene. In conclusion, genistein‐mediated histone modifications of ATF3 may lead to G2 phase arrest and apoptosis in SW620.

MP47-12 TIME COURSE OF APOPTOTIC NEURONAL CELL DEATH IN THE MAJOR PELVIC GANGLION FOLLOWING CAVERNOUS NERVE INJURY

You have accessJournal of UrologySexual Function/Dysfunction/Andrology: Basic Research II1 Apr 2014MP47-12 TIME COURSE OF APOPTOTIC NEURONAL CELL DEATH IN THE MAJOR PELVIC GANGLION FOLLOWING CAVERNOUS NERVE INJURY Johanna Hannan, Maarten Albersen, Joseph Watson, Xiaopu Liu, Petter Hedlund, Arthur Burnett, and Trinity Bivalacqua Johanna HannanJohanna Hannan More articles by this author , Maarten AlbersenMaarten Albersen More articles by this author , Joseph WatsonJoseph Watson More articles by this author , Xiaopu LiuXiaopu Liu More articles by this author , Petter HedlundPetter Hedlund More articles by this author , Arthur BurnettArthur Burnett More articles by this author , and Trinity BivalacquaTrinity Bivalacqua More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2014.02.1465AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail Introduction and Objectives Surgical treatments for prostate, bladder and colorectal cancers often lead to neuronal damage and erectile dysfunction (ED). Despite nerve-sparing techniques, ED prevails and preventing neurodegeneration is of great importance. This study aimed to characterize the markers of neuronal injury and apoptosis in the major pelvic ganglion (MPG) following bilateral cavernous nerve injury (BCNI). Methods Male Sprague-Dawley rats underwent sham or BCNI and MPGs were collected 2, 7, 14, 21, 30 and 60 days following BCNI (n=5/group). MPG gene expression analysis by qPCR was performed for markers of neuronal injury: activating transcription factor 3 (ATF3), glial fibrillary acidic protein (GFAP); apoptotic markers: caspase 1, caspase 3, caspase 9, Bcl2–associated X protein (Bax), BRCA1-associated RING domain protein 1 (BARD1); and neuronal markers: β-tubulin3 (BT3), tyrosine hydroxylase (TH), vesicular acetylcholine transporter (VAChT) and neuronal nitric oxide synthase (nNOS). Additional injured MPGs were fixed, embedded in paraffin and assessed for apoptosis by TUNEL (n=4/group). Results GFAP and ATF3 gene expression were significantly increased in MPGs from 2-21 days following BCNI (p<0.05). Apoptotic markers caspase1, caspase3, Bax and BARD1 at 2-14 days after BCNI were significantly elevated (p<0.05). There was no change in caspase 9 gene expression. Neuronal markers BT3, TH, and VACht were significantly lower in 2 and 7 day injured MPGs (p<0.05). Furthermore, 2 and 7 days following BCNI nNOS gene expression was 50% lower than sham (p<0.05). Apoptotic positive cells were significantly increased in MPGs 7 and 14 days following BCNI (Sham: 0.3±0.12, 7d: 9.5±1.2, 14d: 9.7±3.5, p<0.05). Conclusions Markers of neuronal injury and apoptosis are significantly increased early following BCNI. Further studies inhibiting the apoptotic pathway at these early time points may provide therapeutic avenues for neurogenic-mediated ED. © 2014FiguresReferencesRelatedDetails Volume 191Issue 4SApril 2014Page: e522-e523 Advertisement Copyright & Permissions© 2014MetricsAuthor Information Johanna Hannan More articles by this author Maarten Albersen More articles by this author Joseph Watson More articles by this author Xiaopu Liu More articles by this author Petter Hedlund More articles by this author Arthur Burnett More articles by this author Trinity Bivalacqua More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...