Role Of Activating Transcription Factor 3 Research Articles

ATF3 deficiency promotes alveolar macrophage pyroptosis in sepsis-induced acute lung injury

BackgroundAcute lung injury (ALI) ranks among the leading reasons for death in septic patients. As an essential transcription factor associated with stress, activating transcription factor 3 (ATF3) participates in a variety of pathophysiological processes, including immunology and inflammation. However, the specific mechanism of ATF3 in pyroptosis of sepsis-induced ALI remains elusive. MethodsA mouse model of ALI was established by administering lipopolysaccharide (LPS) in vivo and LPS combined with adenosine triphosphate (ATP) in vitro to compare differences in ATF3 expression level. The role of ATF3 in pyroptosis was then assessed by knocking down ATF3 using small interfering RNA. The levels of interleukin-6 (IL-6), tumor necrosis factor-α, IL-1β, and IL-18 in mouse serum and cell culture supernatants were measured using enzyme-linked immunosorbent assay. Moreover, immunohistochemistry, immunofluorescence, Western blotting, co-immunoprecipitation, and quantitative reverse transcription polymerase chain reaction were employed for examining pyroptosis and pyroptotic pathways. ResultsBoth in vitro and in vivo ALI models were successfully established. LPS could activate the pyroptotic signaling pathway, and the expression level of ATF3 peaked 6h after LPS and ATP stimulation in vitro. ATF3 could interact with NLRP3 and potentially influence the assembly of the inflammasome. This mechanism could involve the inhibition of the classical pyroptotic pathway, including Caspase-1 and Gasdermin D transcription and cleavage, as well as the inhibition of the non-classical pyroptotic pathway, including transcription and cleavage of Caspase-11. ConclusionThe results indicated that inhibition of ATF3 could exacerbate sepsis-induced ALI by regulating pyroptotic pathways. The potential of targeting ATF3 as a future treatment strategy for ALI is noteworthy.

Activating transcription factor 3: A potential therapeutic target for inflammatory pulmonary diseases.

Activating transcription factor 3 (ATF3) is a nuclear protein that is widely expressed in a variety of cells. It is a stress-inducible transcription gene and a member of the activating transcription factor/cAMP responsive element-binding protein (ATF/CREB) family. The comprehensive literature review was conducted by searching PubMed and Google Scholar. Search terms used were "ATF3", "ATF3 and (ALI or ARDS)", "ATF3 and COPD", "ATF3 and PF", and "ATF3 and Posttranslational modifications". Recent studies have shown that ATF3 plays a critical role in many inflammatory pulmonary diseases, including acute lung injury (ALI)/acute respiratory distress syndrome (ARDS), chronic obstructive pulmonary disease (COPD), and pulmonary fibrosis (PF). ATF3 participates in many signaling pathways and complex pathophysiological processes, such as inflammation, immunity, endoplasmic reticulum stress, and cell proliferation. However, the role of ATF3 in current studies is controversial, and there are reports showing that ATF3 plays different roles in different pulmonary diseases. In this review, we first summarized the structure, function, and mechanism of ATF3 in various inflammatory pulmonary diseases. The impact of ATF3 on disease pathogenesis and the clinical implications was particularly focused on, with an overall aim to identify new targets for treating inflammatory pulmonary diseases.

The Role of ATF3 in Neuronal Differentiation and Development of Neuronal Networks in Opossum Postnatal Cortical Cultures.

Activating transcription factor 3 (ATF3), a member of the ATF/cAMP response element-binding (CREB) family, is upregulated by various intracellular and extracellular signals such as injury and signals related to cell proliferation. ATF3 also belongs to the regeneration-associated genes (RAG) group of transcription factors. RAG and ATF/CREB transcription factors that play an important role in embryonic neuronal development and PNS regeneration may also be involved in postnatal neuronal differentiation and development, as well as in the regeneration of the injured CNS. Here we investigated the effect of ATF3 in differentiation, neural outgrowth, network formation, and regeneration after injury using postnatal dissociated cortical neurons derived from neonatal opossums (Monodelphis domestica). Our results show that RAG and ATF genes are differentially expressed in early differentiated neurons versus undifferentiated neurospheres and that many members of those families, ATF3 in particular, are upregulated in cortical cultures obtained from younger animals that have the ability to fully functionally regenerate spinal cord after injury. In addition, we observed different intracellular localization of ATF3 that shifts from nuclear (in neuronal progenitors) to cytoplasmic (in more mature neurons) during neuronal differentiation. The ATF3 inhibition, pharmacological or by specific antibody, reduced the neurite outgrowth and differentiation and caused increased cell death in early differentiating cortical neuronal cultures, suggesting the importance of ATF3 in the CNS development of neonatal opossums. Finally, we investigated the regeneration capacity of primary cortical cultures after mechanical injury using the scratch assay. Remarkably, neonatal opossum-derived cultures retain their capacity to regenerate for up to 1 month in vitro. Inhibition of ATF3 correlates with reduced neurite outgrowth and regeneration after injury. These results indicate that ATF3, and possibly other members of RAG and ATF/CREB family of transcription factors, have an important role both during cortical postnatal development and in response after injury.

Abstract P795: Activating Transcription Factor 3 Improves Stroke Recovery Through Reduction of Neuronal Damage and Neuroinflammation

Background and Purpose: Activating transcription factor 3 (ATF3), a member of the ATF/CREB family, is upregulated in the early phase of various stresses including ischemic stroke. The role of ATF3 in ischemic stroke injury has not been fully elucidated. Hypothesis: ATF3 improves stroke outcomes through reduction of neuronal damage and neuroinflammation. Methods: Permanent distal middle cerebral artery occlusion (pMCAO) was performed in 8-week-old male and female C57BL/6J (WT) and ATF3 knockout (ATF3 -/- ) mice. The sensorimotor functions were analyzed 3 days after pMCAO through adhesive removal and corner tests. Infarct volumes and injured neurons were quantified on Nissl stained and Fluoro-Jade C (FJC) stained sections. The cell-specific expression of ATF3 was analyzed by co-staining ATF3 antibody with antibodies specific to NeuN (Neuron), CD68 (microglia/macrophage) and GFAP (astrocyte). The expression of ATF3 in injured neurons was analyzed by ATF3 and FJC double labeling. Results: ATF3 expression was detected exclusively in neurons in the infarct area 1 day after pMCAO. There were a few ATF3 + CD68 + microglia/macrophages at the peri-infarct regions 2 and 3 days after pMCAO. Almost all FJC + neurons were ATF3 positive. No ATF3 expression was detected in astrocytes. Compared to WT mice, ATF3 -/- mice took longer time to remove the adhesive from their right paws (p<0.001) in adhesive removal test and more left turns in corner test (p<0.001) 3 days after pMCAO. ATF3 -/- mice also had a larger infarct volume (p = 0.014), more FJC + neurons (p=0.002) and CD68 + microglia/macrophages (p=0.003) in the peri-infarct regions than WT mice. Conclusions: Deletion of ATF3 exacerbates neuronal injury, neuroinflammation, and sensorimotor dysfunction in stroke mice, suggesting that upregulation of ATF3 at early stage of stroke improves ischemic stroke recovery through reduction of neuronal damage and neuroinflammation.

Inhibitory role of ATF3 in gastric cancer progression through regulating cell EMT and stemness

BackgroundGastric cancer (GC) is one of the most common cancers and the third leading cause of cancer related mortality worldwide. The 5-year survival rate is rather low owing to advanced unresectable and distant metastasis. The EMT has been widely implicated in the stemness, metastatic dormancy, and chemoresistance of different solid tumors. Given the fact that activating transcription factor-3 (ATF3) is a member of the ATF/CREB family of transcription factors and its role in regulation of GC recurrence and metastasis remain poorly understood, the aim of the present study was to investigate its potential impact in epithelial–mesenchymal transition (EMT) and cancer stem cell (CSC) properties and GC aggression.MethodsTo elucidate the potential role of ATF3 in gastric cancer, we utilized SGC-7901 and MGC-803 gastric cancer cell lines as research models and constructed stable cell lines overexpressing ATF3. We conducted a series of assays including cell proliferation, colony formation, cell migration, tumorsphere formation, and invasion to investigate the functional roles of ATF3 in stemness of gastric cancer. The possible effect of ATF3 on epithelial–mesenchymal transition (EMT) was assessed through flow cytometry and qRT-PCR. In vivo functional effect of upregulation of ATF3 on tumor growth was examined in a mouse xenograft model.ResultsWe found that overexpression of ATF3 inhibited cell proliferation, colony formation, cell migration and invasion. In addition, up-regulation of ATF3 attenuated tumorsphere formation, cell stemness, and potentially decreased expression of EMT markers. Moreover, ATF3 overexpression inhibited tumorigenesis in mouse xenograft model.ConclusionOur data suggest a suppressive role of ATF3 in gastric cancer development. Our findings will provide a potential therapeutic strategy and novel drug target for gastric cancer.

Role of ATF3 as a prognostic biomarker and correlation of ATF3 expression with macrophage infiltration in hepatocellular carcinoma

BackgroundThe abnormal expression of activating transcription factor 3 (ATF3), a member of the basic leucine zipper (bZIP) family of transcription factors, is associated with carcinogenesis. However, the expression pattern and exact role of ATF3 in the development and progression of hepatocellular carcinoma (HCC) remain unclear.MethodsWe used UALCAN, ONCOMINE, Kaplan–Meier plotter, and cBioPortal databases to investigate the prognostic value of ATF3 expression in HCC.ResultsATF3 was found to be expressed at low levels in multiple HCC tumor tissues. Moreover, low ATF3 expression was significantly associated with clinical cancer stage and pathological tumor grade in patients with HCC. Therefore, low expression of ATF3 was significantly associated with poor overall survival in patients with HCC. Functional network analysis showed that ATF3 regulates cytokine receptors and signaling pathways via various cancer-related kinases, miRNAs, and transcription factors. ATF3 expression was found to be correlated with macrophage infiltration levels and with macrophage immune marker sets in HCC patients.ConclusionsUsing data mining methods, we clarified the role of ATF3 expression and related regulatory networks in HCC, laying a foundation for further functional research. Future research will validate our findings and establish clinical applications of ATF3 in the diagnosis and treatment of HCC.

Master Regulator Activating Transcription Factor 3 (ATF3) in Metabolic Homeostasis and Cancer

Activating transcription factor 3 (ATF3) is a stress-induced transcription factor that plays vital roles in modulating metabolism, immunity, and oncogenesis. ATF3 acts as a hub of the cellular adaptive-response network. Multiple extracellular signals, such as endoplasmic reticulum (ER) stress, cytokines, chemokines, and LPS, are connected to ATF3 induction. The function of ATF3 as a regulator of metabolism and immunity has recently sparked intense attention. In this review, we describe how ATF3 can act as both a transcriptional activator and a repressor. We then focus on the role of ATF3 and ATF3-regulated signals in modulating metabolism, immunity, and oncogenesis. The roles of ATF3 in glucose metabolism and adipose tissue regulation are also explored. Next, we summarize how ATF3 regulates immunity and maintains normal host defense. In addition, we elaborate on the roles of ATF3 as a regulator of prostate, breast, colon, lung, and liver cancers. Further understanding of how ATF3 regulates signaling pathways involved in glucose metabolism, adipocyte metabolism, immuno-responsiveness, and oncogenesis in various cancers, including prostate, breast, colon, lung, and liver cancers, is then provided. Finally, we demonstrate that ATF3 acts as a master regulator of metabolic homeostasis and, therefore, may be an appealing target for the treatment of metabolic dyshomeostasis, immune disorders, and various cancers.

The role of ATF3 in ZnO nanoparticle-induced genotoxicity and cytotoxicity in bronchial epithelial cells

ZnO nanoparticle (ZnO NP) exposure causes oxidative stress in the respiratory system, leading to pulmonary damage. Activating transcription factor 3 (ATF3) participates in a variety of cellular stress responses. However, the role of ATF3 in ZnO NP genotoxicity and cytotoxicity remains to be explored. Here we reported that ZnO NP treatment dramatically induced the expression of ATF3 in human bronchial epithelial (HBE) cells, which was mediated by the nuclear factor erythroid 2-related factor 2 (Nrf2). ATF3 was required for the repair of ZnO NP-induced DNA damage as gamma foci number increased when endogenous ATF3 was silenced. Moreover, ATF3 also contributed to ZnO NP-induced cell apoptosis. Mechanistic study revealed that ATF3 interacted with the p53 protein and upregulated its expression under ZnO NP treatment. Collectively, our findings demonstrated ATF3 as an important regulator of epithelial homeostasis by promoting both DNA repair and the death of damaged cells under ZnO NP-induced genotoxic stress.

P 198 - Role of ATF3 in mediating lipid-induced stress signaling in brain microvascular endothelial cells

Elevation of blood triglycerides, primarily triglyceride-rich lipoproteins (TGRL), is an independent risk factor for atherosclerotic cardiovascular disease and Vascular Dementia (VaD). The accumulating evidence indicates that the development of atherosclerosis and VaD are linked to vascular inflammation. Previous studies suggest a relationship of blood lipids and/or lipoproteins to vascular inflammation leading to dementia. The objective of this study is to characterize human brain microvascular endothelial cells (HBMVEC) responses to postprandial TGRL and determine the pathways that have potential to increase risk for vascular dementia. To address this question, we treated HBMVEC with TGRL lipolysis products and then we used NGS-based RNA-Seq methods for transcriptional profiling experiments. Our data demonstrated up-regulation of activating transcription factor 3 (ATF3)-dependent inflammatory pathways in HBMVEC. Additionally, system biology analysis exhibited putative lipid-related targets and corresponding isoforms of ATF3 that may be implicated in hypertriglyceridemia-induced vascular inflammation in brain. Furthermore, RNA-Seq analysis determined specific isoforms of key transcriptional regulator ATF3 and stress response signaling pathway in lipolysis products-mediated brain vascular inflammation which may lead to vascular dementia.

The research progress on the role of ATF3 in the malignant tumor

Activating transcription factor 3 (ATF3) is a protein produced by the cells in the ischemic reperfusion injury, neuronal or liver damage, skin damage, viral oncogene expression or DNA damage. Recent studies have found that ATF3 plays an important role in the occurrence and development of human malignant tumors. Key words: Activating transcription factor 3/ME; Neoplasms/ME; Review

Heart Failure: From Protein to Phenotype37MicroRNA-494 reduces ATF3 expression and promotes heart failure in cardiac hypertrophic remodeling in vivo38A novel recessive plakophilin-2 gene mutation causes severe arrhythmogenic dilated cardiomyopathy and sudden cardiac death at young age39Investigation of titin expression in explanted hearts with familial dilated cardiomyopathy and TTN truncating variants

Heart Failure: From Protein to Phenotype37MicroRNA-494 reduces ATF3 expression and promotes heart failure in cardiac hypertrophic remodeling in vivo38A novel recessive plakophilin-2 gene mutation causes severe arrhythmogenic dilated cardiomyopathy and sudden cardiac death at young age39Investigation of titin expression in explanted hearts with familial dilated cardiomyopathy and TTN truncating variants

The Stress-responsive Gene ATF3 Mediates Dichotomous UV Responses by Regulating the Tip60 and p53 Proteins

The response to UV irradiation is important for a cell to maintain its genetic integrity when challenged by environmental genotoxins. An immediate early response to UV irradiation is the rapid induction of activating transcription factor 3 (ATF3) expression. Although emerging evidence has linked ATF3 to stress pathways regulated by the tumor suppressor p53 and the histone acetyltransferase Tip60, the role of ATF3 in the UV response remains largely unclear. Here, we report that ATF3 mediated dichotomous UV responses. Although UV irradiation enhanced the binding of ATF3 to Tip60, knockdown of ATF3 expression decreased Tip60 stability, thereby impairing Tip60 induction by UV irradiation. In line with the role of Tip60 in mediating UV-induced apoptosis, ATF3 promoted the death of p53-defective cells in response to UV irradiation. However, ATF3 could also activate p53 and promote p53-mediated DNA repair, mainly through altering histone modifications that could facilitate recruitment of DNA repair proteins (such as DDB2) to damaged DNA sites. As a result, ATF3 rather protected the p53 wild-type cells from UV-induced apoptosis. Our results thus indicate that ATF3 regulates cell fates upon UV irradiation in a p53-dependent manner.

ATF3 provides protection from Staphylococcus aureus and Listeria monocytogenes infections.

Activating transcription factor 3 (ATF3) is a stress-induced transcriptional regulator in eukaryote. The role of ATF3 in cancer has been well defined, but how ATF3 functions in bacterial infection is not well understood. Pneumococcal infection has been shown to induce ATF3 expression, which subsequently enhances cytokine production and provides protection from lethal Streptococcus pneumoniae infection, but the role of ATF3 in other Gram-positive (G(+)) infections remains unclear. Here, we report that infection with other G(+) bacteria (Staphylococcus aureus and Listeria monocytogenes) and with G(-) bacteria (uropathogenic Escherichia coli) also significantly induced ATF3 expression. Moreover, the production of cytokines (tumor necrosis factor alpha [TNF]-α, interleukin [IL]-1β, IL-6 and interferon [IFN]-γ) was enhanced by ATF3 in S.aureus and L. monocytogenes infection, but decreased in uropathogenic E. coli (UPEC) infection. In addition, in S.aureus and L. monocytogenes infections, ATF3 WT mice cleared bacteria more efficiently and had higher survival rates than ATF3 knockout mice. However, in UPEC infection, no significant difference was found in survival rate. Taken together, these data suggest that ATF3 provides protection from S.aureus and L. monocytogenes infections; however, the role of ATF3 in UPEC infection is more complicated and should be further elucidated.

Loss of ATF3 promotes Akt activation and prostate cancer development in a Pten knockout mouse model.

Activating transcription factor 3 (ATF3) responds to diverse cellular stresses, and regulates oncogenic activities (e.g., proliferation, survival and migration) through direct transcriptional regulation or protein-protein interactions. Although aberrant ATF3 expression is frequently found in human cancers, the role of ATF3 in tumorigenesis is poorly understood. Here we demonstrate that ATF3 suppresses the development of prostate cancer induced by knockout of the tumor suppressor Pten in mouse prostates. Where as the oncogenic stress elicited by Pten loss induced ATF3 expression in prostate epithelium, we found that ATF3 deficiency increased cell proliferation and promoted cell survival, leading to early onset of mouse prostatic intraepithelial neoplasia and the progression of prostate lesions to invasive adenocarcinoma. Importantly, loss of ATF3 promoted activation of the oncogenic AKT signaling evidenced by high levels of phosphorylated AKT and S6 proteins in ATF3-null prostate lesions. In line with these in vivo results, knockdown of ATF3 expression in human prostate cancer cells by sgRNA-mediated targeting activated AKT and increased matrix metalloproteinase-9 expression. Our results thus link ATF3 to the AKT signaling, and suggest that ATF3 is a tumor suppressor for the major subset of prostate cancers harboring dysfunctional Pten.

Role of Activating Transcription Factor 3 (ATF3) in Endoplasmic Reticulum (ER) Stress-induced Sensitization of p53-deficient Human Colon Cancer Cells to Tumor Necrosis Factor (TNF)-related Apoptosis-inducing Ligand (TRAIL)-mediated Apoptosis through Up-regulation of Death Receptor 5 (DR5) by Zerumbone and Celecoxib

Death receptor 5 (DR5) is a death domain-containing transmembrane receptor that triggers cell death upon binding to its ligand, TNF-related apoptosis-inducing ligand (TRAIL), and a combination of TRAIL and agents that increase the expression of DR5 is expected to be a novel anticancer therapy. In this report, we demonstrate that the stress response gene ATF3 is required for endoplasmic reticulum stress-mediated DR5 induction upon zerumbone (ZER) and celecoxib (CCB) in human p53-deficient colorectal cancer cells. Both agents activated PERK-eIF2α kinases and induced the expression of activating transcription factor 4 (ATF4)-CCAAT enhancer-binding protein (C/EBP) homologous protein, which were remarkably suppressed by reactive oxygen species scavengers. In the absence of ATF3, the induction of DR5 mRNA and protein was abrogated significantly, and this was associated with reduced cell death by cotreatment of TRAIL with ZER or CCB. By contrast, exogenous expression of ATF3 caused a more rapid and elevated expression of DR5, resulting in enhanced sensitivity to apoptotic cell death by TRAIL/ZER or TRAIL/CCB. A reporter assay demonstrated that at least two ATF/cAMP response element motifs as well as C/EBP homologous protein motif at the proximal region of the human DR5 gene promoter were required for ZER-induced DR5 gene transcription. Taken together, our results provide novel insights into the role of ATF3 as an essential transcription factor for p53-independent DR5 induction upon both ZER and CCB treatment, and this may be a useful biomarker for TRAIL-based anticancer therapy.

Activating transcription factor 3 promotes colon cancer metastasis

Colorectal cancer is one of the commonest of solid malignancy in the world. Activating transcription factor 3 (ATF3), a homolog of the mouse TI-241 and rat LFR-1, is a stress responsive gene that has been widely indicated in different malignancies. However, the role of ATF3 in colon cancer is paradoxical with both a suggested pro- and anti-tumorigenic role. The objective of the current study was to investigate the role of ATF3 in colon cancer metastasis using HT29 and CaCO2 colon cancer cell lines. Expression of ATF3 was initially evaluated in five pairs of colon cancer and matched noncancerous colon tissues. The role of ATF3 in promoting in vitro migration and invasion were evaluated by siRNA-mediated knockdown and adenovirus-mediated overexpression of ATF3. In addition, the role of ATF3 in promoting in vivo tumor growth and hepatic metastasis was investigated by shRNA-mediated knockdown of ATF3. Expression of ATF3 was more in the colon cancer tissues as compared with the pooled noncancerous control colon tissue. Our results showed that in both HT29 and CaCO2 cells, ATF3 promoted in vitro motility and invasion. Furthermore, knockdown of ATF3 attenuated subcutaneous tumor growth and CD31(+) neovasculature in xenograft assays with HT29 and CaCO2 cells and inhibited hepatic metastasis. Cumulatively, our results unequivocally show that ATF3 promotes colon cancer metastasis.

Activating Transcription Factor 3 Contributes to Toll-Like Receptor-Mediated Macrophage Survival via Repression ofBaxandBak

Macrophages play an essential role in the innate immune response to infection and tissue injury. However, excessive macrophage activation may also significantly contribute to chronic inflammatory diseases. The Toll-like receptor (TLR) family are key regulators of innate immune responses in macrophages, and they are able to promote their survival and resistance against apoptosis. We, and others, have shown that the adaptive response gene, activating transcription factor 3 (ATF3), acts as a negative regulator of TLR signaling by repressing transcription of pro-inflammatory cytokines in primary mouse macrophages. Here, we describe a novel role for ATF3 as a component of TLR-mediated survival in macrophages. ATF3-deficient bone marrow macrophages show reduced survival in response to a range of TLR ligands and significantly higher apoptotic rates were observed in response to lipopolysaccharide, indicating that ATF3 is required to suppress apoptosis in macrophages. Furthermore, we show that ATF3 lies downstream of JNK signaling after TLR engagement, resulting in repression of pro-apoptotic Bak and Bax transcription.

Abstract 5442: The role of ATF3 in arsenic-induced GADD45 expression.

Abstract GADD45 (growth arrest and DNA damage inducible gene 45), an effector of G2/M checkpoint, is induced by trivalent arsenic (As3+) in human bronchial epithelial cell line, BEAS-2B. In the present study, we investigated the regulation of ATF3 (activating transcription factor 3) on GADD45 with As 3+ treatment. We demonstrated that As3+ induced ATF3 expression, at both mRNA and protein level, and the induction was reduced by p38 and JNK inhibition, but not affected by PI3K and ERK inhibition. Luciferase assay also confirmed As3+ involvement in ATF3 transcriptional regulation. Moreover, silencing of ATF3 downregulated As3+-induced expression of GADD45 and chromatin immunoprecipitation suggested that ATF3 acted as an activator of GADD45 transcription in the presence of As3+. Taken together, our data indicated that the increase of GADD45 in response to As3+ was mediated by ATF3. Citation Format: Qiwen Shi, Deepak Bhatia, Vijaykumar B. Sutariya. The role of ATF3 in arsenic-induced GADD45 expression. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5442. doi:10.1158/1538-7445.AM2013-5442

Abstract 895: Activating transcription factor 3 (ATF3) down-regulation correlates with platinum resistance in non-small cell lung cancer (NSCLC).

Abstract Background: NSCLC is the most common cause of cancer-related death. Platinum-based chemotherapy is the mainstay of treatment, but a variety of mechanisms lead to platinum-resistance. ATF3 is a transcription factor, activated in response to a wide variety of stress signals including DNA damage and hypoxia. We recently demonstrated a role for ATF3 as an important regulator of platinum-induced cytotoxicity. In this study, we hypothesize that ATF3 expression correlates with platinum-sensitivity/resistance in NSCLC. Methods: ATF3 induction was examined by Western blots and real-time RT-PCR in isogenic sets of platinum- sensitive (S) or induced resistant (R) NSCLC cell lines. A 1200 compound library was screened to identify platinum-sensitizers in both sets of cell lines. Complete RNA sequencing (RNA-seq) was performed in parental (S) and resistant (R) derived cell lines comparing expression patterns of either untreated or platinum treated cells. Similarly, platinum R and S tumors were identified by screening NSCLC patients’ clinical records and expression patterns compared with RNA-seq analysis. Results: Both mRNA and potein levels of ATF3 were induced 35-120 fold (mRNA) by cisplatin treatment in S cell lines, but only 1-12 fold (mRNA) in the corresponding R lines. The 1200 compound library screen of FDA approved compounds identified several commonly used chemotherapeutic agents as sensitizers of platinum cytotoxicity, as well as vorinostat, a histone deacetylase inhibitor that sensitized only S but not R cell lines. An analogue of this agent, M344, enhanced ATF3 induction and cisplatin cytotoxicity in the S lines but not in the R lines. Comparing RNA-seq results of the S and R lines revealed up-regulation of GADD45A, ATF3 and DDIT3/CHOP in the S but not R cell lines. Comparing samples of S and R NSCLC tumors by RNA-seq demonstrated ATF3 to be among the five genes significantly up-regulated in the S tumors compared to the R tumors. Conclusions: Stress-induced ATF3 is correlated with platinum-sensitivity in NSCLC cells. Tumor ATF3 levels represent a potential predictive marker of response to platinum-based treatment in NSCLC. Revealing the mechanism of ATF3 induction by platinum may lead to the identification of novel platinum-sensitizing therapeutic targets. Citation Format: Jair Bar, Ivan Gorn-Hondermann, Reid Stephanie, Patricia Moretto, Iris Shiran, Shlomit Jessel, Marina Perelman, Eyal Heller, Iris Kamer, Inbal Daniel-Meshulam, Glenwood D. Goss, Jim Dimitroulakos. Activating transcription factor 3 (ATF3) down-regulation correlates with platinum resistance in non-small cell lung cancer (NSCLC). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 895. doi:10.1158/1538-7445.AM2013-895

Microarray Analysis of Altered Gene Expression and the Role of ATF3 in HK-2 Cells Treated with Hemin

Objective: To identify gene expression changes and the role of activating transcription factor 3 (ATF3) in hemin toxicity in renal tubular epithelial cells, then elucidate molecular mechanisms of hemin toxicity on renal tubular epithelial cells.Methods: An oligo array comprising 35,035 genes was used to compare differential gene expression in hemin-treated and non-treated HK-2 cells (human renal proximal tubular epithelial cells), and the role of ATF3 in hemin toxicity was assessed using siRNA technique.Results: A total of 128 mRNAs were at least twofold up-regulated and 101 mRNAs were at least twofold down-regulated after hemin treatment. Expression levels of ATF3, heat shock protein 70, c-fos, and c-jun were remarkably increased. Hemin also suppressed nuclear factor-kappa B inhibitor α, β-2 adrenergic receptor, and interleukin-6 mRNA amounts more than twofold. We further demonstrated the protective role of ATF3 in hemin cytotoxicity.Conclusions: The data suggest that hemin caused multiple changes of gene expression in HK-2 cells, and ATF3 protects against hemin cytotoxicity.