Electrophilic properties of itaconate and derivatives regulate the IκBζ-ATF3 inflammatory axis.

Metabolic regulation has been recognized as a powerful principle guiding immune responses. Inflammatory macrophages undergo extensive metabolic rewiring 1 marked by the production of substantial amounts of itaconate, which has recently been described as an immunoregulatory metabolite 2 . Itaconate and its membrane-permeable derivative dimethyl itaconate (DI) selectively inhibit a subset of cytokines 2 , including IL-6 and IL-12 but not TNF. The major effects of itaconate on cellular metabolism during macrophage activation have been attributed to the inhibition of succinate dehydrogenase2,3, yet this inhibition alone is not sufficient to account for the pronounced immunoregulatory effects observed in the case of DI. Furthermore, the regulatory pathway responsible for such selective effects of itaconate and DI on the inflammatory program has not been defined. Here we show that itaconate and DI induce electrophilic stress, react with glutathione and subsequently induce both Nrf2 (also known as NFE2L2)-dependent and -independent responses. We find that electrophilic stress can selectively regulate secondary, but not primary, transcriptional responses to toll-like receptor stimulation via inhibition of IκBζ protein induction. The regulation of IκBζ is independent of Nrf2, and we identify ATF3 as its key mediator. The inhibitory effect is conserved across species and cell types, and the in vivo administration of DI can ameliorate IL-17-IκBζ-driven skin pathology in a mouse model of psoriasis, highlighting the therapeutic potential of this regulatory pathway. Our results demonstrate that targeting the DI-IκBζ regulatory axis could be an important new strategy for the treatment of IL-17-IκBζ-mediated autoimmune diseases.

Chemotherapy-Induced Changes in the Lung Microenvironment: The Role of MMP-2 in Facilitating Intravascular Arrest of Breast Cancer Cells.

Previously, we showed that mice treated with cyclophosphamide (CTX) 4 days before intravenous injection of breast cancer cells had more cancer cells in the lung at 3 h after cancer injection than control counterparts without CTX. At 4 days after its injection, CTX is already excreted from the mice, allowing this pre-treatment design to reveal how CTX may modify the lung environment to indirectly affect cancer cells. In this study, we tested the hypothesis that the increase in cancer cell abundance at 3 h by CTX is due to an increase in the adhesiveness of vascular wall for cancer cells. Our data from protein array analysis and inhibition approach combined with in vitro and in vivo assays support the following two-prong mechanism. (1) CTX increases vascular permeability, resulting in the exposure of the basement membrane (BM). (2) CTX increases the level of matrix metalloproteinase-2 (MMP-2) in mouse serum, which remodels the BM and is functionally important for CTX to increase cancer abundance at this early stage. The combined effect of these two processes is the increased accessibility of critical protein domains in the BM, resulting in higher vascular adhesiveness for cancer cells to adhere. The critical protein domains in the vascular microenvironment are RGD and YISGR domains, whose known binding partners on cancer cells are integrin dimers and laminin receptor, respectively.

Stress-Inducible Gene Atf3 Dictates a Dichotomous Macrophage Activity in Chemotherapy-Enhanced Lung Colonization.

Previously, we showed that chemotherapy paradoxically exacerbated cancer cell colonization at the secondary site in a manner dependent on Atf3, a stress-inducible gene, in the non-cancer host cells. Here, we present evidence that this phenotype is established at an early stage of colonization within days of cancer cell arrival. Using mouse breast cancer models, we showed that, in the wild-type (WT) lung, cyclophosphamide (CTX) increased the ability of the lung to retain cancer cells in the vascular bed. Although CTX did not change the WT lung to affect cancer cell extravasation or proliferation, it changed the lung macrophage to be pro-cancer, protecting cancer cells from death. This, combined with the initial increase in cell retention, resulted in higher lung colonization in CTX-treated than control-treated mice. In the Atf3 knockout (KO) lung, CTX also increased the ability of lung to retain cancer cells. However, the CTX-treated KO macrophage was highly cytotoxic to cancer cells, resulting in no increase in lung colonization-despite the initial increase in cell retention. In summary, the status of Atf3 dictates the dichotomous activity of macrophage: pro-cancer for CTX-treated WT macrophage but anti-cancer for the KO counterpart. This dichotomy provides a mechanistic explanation for CTX to exacerbate lung colonization in the WT but not Atf3 KO lung.

Stress-inducible gene Atf3 in the noncancer host cells contributes to chemotherapy-exacerbated breast cancer metastasis.

Chemotherapy is a double-edged sword. It is anticancer because of its cytotoxicity. Paradoxically, by increasing chemoresistance and cancer metastasis, it is also procancer. However, the underlying mechanisms for chemotherapy-induced procancer activities are not well understood. Here we describe the ability of paclitaxel (PTX), a frontline chemotherapeutic agent, to exacerbate metastasis in mouse models of breast cancer. We demonstrate that, despite the apparent benefit of reducing tumor size, PTX increased the circulating tumor cells in the blood and enhanced the metastatic burden at the lung. At the primary tumor, PTX increased the abundance of the tumor microenvironment of metastasis, a landmark microanatomical structure at the microvasculature where cancer cells enter the blood stream. At the metastatic lung, PTX improved the tissue microenvironment (the "soil") for cancer cells (the "seeds") to thrive; these changes include increased inflammatory monocytes and reduced cytotoxicity. Importantly, these changes in the primary tumor and the metastatic lung were all dependent on Atf3, a stress-inducible gene, in the noncancer host cells. Together, our data provide mechanistic insights into the procancer effect of chemotherapy, explaining its paradox in the context of the seed-and-soil theory. Analyses of public datasets suggest that our data may have relevance to human cancers. Thus, ATF3 in the host cells links a chemotherapeutic agent-a stressor-to immune modulation and cancer metastasis. Dampening the effect of ATF3 may improve the efficacy of chemotherapy.

Activating transcription factor 3 attenuates chemokine and cytokine expression in mouse skeletal muscle after exercise and facilitates molecular adaptation to endurance training.

Activating transcription factor (ATF)3 regulates the expression of inflammation-related genes in several tissues under pathological contexts. In skeletal muscle, atf3 expression increases after exercise, but its target genes remain unknown. We aimed to identify those genes and to determine the influence of ATF3 on muscle adaptation to training. Skeletal muscles of ATF3-knockout (ATF3-KO) and control mice were analyzed at rest, after exercise, and after training. In resting muscles, there was no difference between genotypes in enzymatic activities or fiber type. After exercise, a microarray analysis in quadriceps revealed ATF3 affects genes modulating chemotaxis and chemokine/cytokine activity. Quantitative PCR showed that the mRNA levels of chemokine C-C motif ligand (ccl)8 and chemokine C-X-C motif ligand (cxcl)13 were higher in quadriceps of ATF3-KO mice than in control mice. The same was observed for ccl9 and cxcl13 in soleus. Also in soleus, ccl2, interleukin (il)6, il1ß, and cluster of differentiation (cd)68 mRNA levels increased after exercise only in ATF3-KO mice. Endurance training increased the basal mRNA level of hexokinase-2, hormone sensitive lipase, glutathione peroxidase-1, and myosin heavy chain IIa in quadriceps of control mice but not in ATF3-KO mice. In summary, ATF3 attenuates the expression of inflammation-related genes after exercise and thus facilitates molecular adaptation to training.-Fernández-Verdejo, R., Vanwynsberghe, A. M., Essaghir, A., Demoulin, J.-B., Hai, T., Deldicque, L., Francaux, M. Activating transcription factor 3 attenuates chemokine and cytokine expression in mouse skeletal muscle after exercise and facilitates molecular adaptation to endurance training.

ATF3 mediates inhibitory effects of ethanol on hepatic gluconeogenesis.

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.

Chemotherapy-Exacerbated Breast Cancer Metastasis: A Paradox Explainable by Dysregulated Adaptive-Response.

An emerging picture in cancer biology is that, paradoxically, chemotherapy can actively induce changes that favor cancer progression. These pro-cancer changes can be either inside (intrinsic) or outside (extrinsic) the cancer cells. In this review, we will discuss the extrinsic pro-cancer effect of chemotherapy; that is, the effect of chemotherapy on the non-cancer host cells to promote cancer progression. We will focus on metastasis, and will first discuss recent data from mouse models of breast cancer. Despite reducing the size of primary tumors, chemotherapy changes the tumor microenvironment, resulting in an increased escape of cancer cells into the blood stream. Furthermore, chemotherapry changes the tissue microenvironment at the distant sites, making it more hospitable to cancer cells upon their arrival. We will then discuss the idea and evidence that these devastating pro-metastatic effects of chemotherapy can be explained in the context of adaptive-response. At the end, we will discuss the potential relevance of these mouse data to human breast cancer and their implication on chemotherapy in the clinic.

Activating transcription factor 3 regulates canonical TGFß signalling in systemic sclerosis.

BACKGROUND: Activating transcription factor 3 (ATF3), a member of the ATF/cAMP-responsive element binding (CREB) family of transcription factors, regulates cellular response to stress including oxidative stress. The aim of this study was to analyse the role of ATF3 in fibroblast activation in systemic sclerosis (SSc). METHODS: ATF3 was analysed by reverse transcription quantitative PCR, western blot and immunohistochemistry. ATF3 knockout fibroblasts and mice were used to study the functional role of ATF3. Knockdown experiments, reporter assays and coimmunoprecipitation were performed to study the effects of ATF3 on Smad and activation protein 1 (AP-1) signalling. The role of c-Jun was analysed by costaining, specific inactivation and coimmunoprecipitation. RESULTS: Transforming growth factor-ß (TGFß) upregulates the expression of ATF3 in SSc fibroblasts. ATF3-deficient fibroblasts were less sensitive to TGFß, whereas ectopic expression of ATF3 enhanced the profibrotic effects of TGFß. Mechanistically, ATF3 interacts with Smad3 directly on stimulation with TGFß and regulates Smad activity in a c-Jun-dependent manner. Knockout of ATF3 protected mice from bleomycin-induced fibrosis and fibrosis induced by overexpression of a constitutively active TGFß receptor I. Reporter assays and analyses of the expression of Smad target genes demonstrated that binding of ATF3 regulates the transcriptional activity of Smad3. CONCLUSIONS: We demonstrate for the first time a key role for ATF3 in fibrosis. Knockout of the ATF3 gene reduced the stimulatory effect of TGFß on fibroblasts by interfering with canonical Smad signalling and protected the mice from experimental fibrosis in two different models. ATF3 might thus be a candidate for molecular targeted therapies for SSc.

ATF3 is a novel regulator of mouse neutrophil migration.

Expression of the activating transcription factor 3 (ATF3) gene is induced by Toll-like receptor (TLR) signaling. In turn, ATF3 protein inhibits the expression of various TLR-driven proinflammatory genes. Given its counter-regulatory role in diverse innate immune responses, we defined the effects of ATF3 on neutrophilic airway inflammation in mice. ATF3 deletion was associated with increased lipopolysaccharide (LPS)-driven airway epithelia production of CXCL1, but not CXCL2, findings concordant with a consensus ATF3-binding site identified solely in the Cxcl1 promoter. Unexpectedly, ATF3-deficient mice did not exhibit increased airway neutrophilia after LPS challenge. Bone marrow chimeras revealed a specific reduction in ATF3(-/-) neutrophil recruitment to wild-type lungs. In vitro, ATF3(-/-) neutrophils exhibited a profound chemotaxis defect. Global gene expression analysis identified ablated Tiam2 expression in ATF3(-/-) neutrophils. TIAM2 regulates cellular motility by activating Rac1-mediated focal adhesion disassembly. Notably, ATF3(-/-) and ATF3-sufficient TIAM2 knockdown neutrophils, both lacking TIAM2, exhibited increased focal complex area, along with excessive CD11b-mediated F-actin polymerization. Together, our data describe a dichotomous role for ATF3-mediated regulation of neutrophilic responses: inhibition of neutrophil chemokine production but promotion of neutrophil chemotaxis.

ßIV-Spectrin and CaMKII facilitate Kir6.2 regulation in pancreatic beta cells.

Identified over a dozen years ago in the brain and pancreatic islet, ßIV-spectrin is critical for the local organization of protein complexes throughout the nervous system. ßIV-Spectrin targets ion channels and adapter proteins to axon initial segments and nodes of Ranvier in neurons, and ßIV-spectrin dysfunction underlies ataxia and early death in mice. Despite advances in ßIV-spectrin research in the nervous system, its role in pancreatic islet biology is unknown. Here, we report that ßIV-spectrin serves as a multifunctional structural and signaling platform in the pancreatic islet. We report that ßIV-spectrin directly associates with and targets the calcium/calmodulin-dependent protein kinase II (CaMKII) in pancreatic islets. In parallel, ßIV-spectrin targets ankyrin-B and the ATP-sensitive potassium channel. Consistent with these findings, ßIV-spectrin mutant mice lacking CaMKII- or ankyrin-binding motifs display selective loss of expression and targeting of key protein components, including CaMKIIδ. ßIV-Spectrin-targeted CaMKII directly phosphorylates the inwardly-rectifying potassium channel, Kir6.2 (alpha subunit of KATP channel complex), and we identify the specific residue, Kir6.2 T224, responsible for CaMKII-dependent regulation of KATP channel function. CaMKII-dependent phosphorylation alters channel regulation resulting in KATP channel inhibition, a cellular phenotype consistent with aberrant insulin regulation. Finally, we demonstrate aberrant KATP channel phosphorylation in ßIV-spectrin mutant mice. In summary, our findings establish a broader role for ßIV-spectrin in regulation of cell membrane excitability in the pancreatic islet, define the pathway for CaMKII local control in pancreatic beta cells, and identify the mechanism for CaMKII-dependent regulation of KATP channels.

Atf3 negatively regulates Ptgs2/Cox2 expression during acute inflammation.

By generating prostaglandins, cyclooxygenase-2 (Cox-2/Ptgs2) plays a critical role in regulating inflammatory responses. While several inflammatory stimuli have been shown to increase Ptgs2 expression, less is known about how the transcription of this gene is terminated. Here we show that stimulation of macrophages with yeast zymosan, a TLR2/6 and dectin-1 agonist, causes a transient increase in the expression of Ptgs2 accompanied by a simultaneous increase in the expression of the transcriptional repressor, activating transcription factor-3 (Atf3). The expression of Ptgs2 was significantly higher in resident peritoneal macrophages isolated from Atf3(-/-) mice than that from Atf3(+/+) mice and was associated with higher prostaglandin production upon stimulation with zymosan. In activated macrophages, Atf3 accumulated in the nucleus and chromatin-immunoprecipitation analysis showed that Atf3 is recruited to the Ptgs2 promoter region. In acute peritonitis and in cutaneous wounds, there was increased leukocyte accumulation and higher levels of prostaglandins (PGE2/PGD2) in inflammatory exudates of Atf3(-/-) mice compared with WT mice. Collectively, these results demonstrate that during acute inflammation Atf3 negatively regulates Ptgs2 and therefore dysregulation of this axis could potentially contribute to aberrant Ptgs2 expression in chronic inflammatory diseases. Moreover, this axis could be a new therapeutic target for suppressing Ptgs2 expression and the resultant inflammatory responses.

The bZIP repressor proteins, c-Jun dimerization protein 2 and activating transcription factor 3, recruit multiple HDAC members to the ATF3 promoter.

JDP2, is a basic leucine zipper (bZIP) protein displaying a high degree of homology with the stress inducible transcription factor, ATF3. Both proteins bind to cAMP and TPA response elements and repress transcription by multiple mechanisms. Histone deacetylases (HDACs) play a key role in gene inactivation by deacetylating lysine residues on histones. Here we describe the association of JDP2 and ATF3 with HDACs 1, 2-6 and 10. Association of HDAC3 and HDAC6 with JDP2 and ATF3 occurs via direct protein-protein interactions. Only part of the N-terminal bZIP motif of JDP2 and ATF3 basic domain is necessary and sufficient for the interaction with HDACs in a manner that is independent of coiled-coil dimerization. Class I HDACs associate with the bZIP repressors via the DAC conserved domain whereas the Class IIb HDAC6 associates through its C-terminal unique binder of ubiquitin Zn finger domain. Both JDP2 and ATF3 are known to bind and repress the ATF3 promoter. MEF cells treated with histone deacetylase inhibitor, trichostatin A (TSA) display enhanced ATF3 transcription. ATF3 enhanced transcription is significantly reduced in MEF cells lacking both ATF3 and JDP2. Collectively, we propose that the recruitment of multiple HDAC members to JDP2 and ATF3 is part of their transcription repression mechanism.

ATF3, an adaptive-response gene, enhances TGF{beta} signaling and cancer-initiating cell features in breast cancer cells.

The activating transcription factor 3 (ATF3) gene is induced by a variety of signals, including many of those encountered by cancer cells. We present evidence that ATF3 is induced by TGFß in the MCF10CA1a breast cancer cells and plays an integral role for TGFß to upregulate its target genes snail, slug and twist, and to enhance cell motility. Furthermore, ATF3 upregulates the expression of the TGFb gene itself, forming a positive-feedback loop for TGFß signaling. Functionally, ectopic expression of ATF3 leads to morphological changes and alterations of markers consistent with epithelial-to-mesenchymal transition (EMT). It also leads to features associated with breast-cancer-initiating cells: increased CD24(low)-CD44(high) population of cells, mammosphere formation and tumorigenesis. Conversely, knockdown of ATF3 reduces EMT, CD24(low)-CD44(high) cells and mammosphere formation. Importantly, knocking down twist, a downstream target, reduces the ability of ATF3 to enhance mammosphere formation, indicating the functional significance of twist in ATF3 action. To our knowledge, this is the first report demonstrating the ability of ATF3 to enhance breast cancer-initiating cell features and to feedback on TGFß. Because ATF3 is an adaptive-response gene and is induced by various stromal signals, these findings have significant implications for how the tumor microenvironment might affect cancer development.

Heat shock transcription factor 1 inhibits expression of IL-6 through activating transcription factor 3.

The febrile response is a complex physiological reaction to disease, including a cytokine-mediated increase in body temperature and the activation of inflammatory systems. Fever has beneficial roles in terms of disease prognosis, partly by suppressing the expression of inflammatory cytokines. However, the molecular mechanisms underlining the fever-mediated suppression of inflammatory gene expression have not been clarified. In this study, we showed that heat shock suppresses LPS-induced expression of IL-6, a major pyrogenic cytokine, in mouse embryonic fibroblasts and macrophages. Heat shock transcription factor 1 (HSF1) activated by heat shock induced the expression of activating transcription factor (ATF) 3, a negative regulator of IL-6, and ATF3 was necessary for heat-mediated suppression of IL-6, indicating a fever-mediated feedback loop consisting of HSF1 and ATF3. A comprehensive analysis of inflammatory gene expression revealed that heat pretreatment suppresses LPS-induced expression of most genes (86%), in part (67%) via ATF3. When HSF1-null and ATF3-null mice were injected with LPS, they expressed much higher levels of IL-6 than wild-type mice, resulting in an exaggerated febrile response. These results demonstrate a novel inhibitory pathway for inflammatory cytokines.

Systems biology approaches identify ATF3 as a negative regulator of Toll-like receptor 4.

The innate immune system is absolutely required for host defence, but, uncontrolled, it leads to inflammatory disease. This control is mediated, in part, by cytokines that are secreted by macrophages. Immune regulation is extraordinarily complex, and can be best investigated with systems approaches (that is, using computational tools to predict regulatory networks arising from global, high-throughput data sets). Here we use cluster analysis of a comprehensive set of transcriptomic data derived from Toll-like receptor (TLR)-activated macrophages to identify a prominent group of genes that appear to be regulated by activating transcription factor 3 (ATF3), a member of the CREB/ATF family of transcription factors. Network analysis predicted that ATF3 is part of a transcriptional complex that also contains members of the nuclear factor (NF)-kappaB family of transcription factors. Promoter analysis of the putative ATF3-regulated gene cluster demonstrated an over-representation of closely apposed ATF3 and NF-kappaB binding sites, which was verified by chromatin immunoprecipitation and hybridization to a DNA microarray. This cluster included important cytokines such as interleukin (IL)-6 and IL-12b. ATF3 and Rel (a component of NF-kappaB) were shown to bind to the regulatory regions of these genes upon macrophage activation. A kinetic model of Il6 and Il12b messenger RNA expression as a function of ATF3 and NF-kappaB promoter binding predicted that ATF3 is a negative regulator of Il6 and Il12b transcription, and this hypothesis was validated using Atf3-null mice. ATF3 seems to inhibit Il6 and Il12b transcription by altering chromatin structure, thereby restricting access to transcription factors. Because ATF3 is itself induced by lipopolysaccharide, it seems to regulate TLR-stimulated inflammatory responses as part of a negative-feedback loop.

ERAD inhibitors integrate ER stress with an epigenetic mechanism to activate BH3-only protein NOXA in cancer cells.

The ubiquitin-proteasome system has recently emerged as a major target for drug development in cancer therapy. The proteasome inhibitor bortezomib has clinical activity in multiple myeloma and mantle cell lymphoma. Here we report that Eeyarestatin I (EerI), a chemical inhibitor that blocks endoplasmic reticulum (ER)-associated protein degradation, has antitumor and biologic activities similar to bortezomib and can synergize with bortezomib. Like bortezomib, EerI-induced cytotoxicity requires the up-regulation of the Bcl-2 homology3 (BH3)-only pro-apoptotic protein NOXA. We further demonstrate that both EerI and bortezomib activate NOXA via an unanticipated mechanism that requires cooperation between two processes. First, these agents elicit an integrated stress response program at the ER to activate the CREB/ATF transcription factors ATF3 and ATF4. We show that ATF3 and ATF4 form a complex capable of binding to the NOXA promoter, which is required for NOXA activation. Second, EerI and bortezomib also block ubiquitination of histone H2A to relieve its inhibition on NOXA transcription. Our results identify a class of anticancer agents that integrate ER stress response with an epigenetic mechanism to induce cell death.

Tumor factors stimulate lysosomal degradation of tumor antigens and undermine their cross-presentation in lung cancer.

Activities of dendritic cells (DCs) that present tumor antigens are often suppressed in tumors. Here we report that this suppression is induced by tumor microenvironment-derived factors, which activate the activating transcription factor-3 (ATF3) transcription factor and downregulate cholesterol 25-hydroxylase (CH25H). Loss of CH25H in antigen presenting cells isolated from human lung tumors is associated with tumor growth and lung cancer progression. Accordingly, mice lacking CH25H in DCs exhibit an accelerated tumor growth, decreased infiltration and impaired activation of intratumoral CD8+ T cells. These mice do not establish measurable long-term immunity against malignant cells that undergo chemotherapy-induced immunogenic cell death. Mechanistically, downregulation of CH25H stimulates membrane fusion between endo-phagosomes and lysosomes, accelerates lysosomal degradation and restricts cross-presentation of tumor antigens in the intratumoral DCs. Administration of STING agonist MSA-2 reduces the lysosomal activity in DCs, restores antigen cross presentation, and increases therapeutic efficacy of PD-1 blockade against tumour challenge in a CH25H-dependent manner. These studies highlight the importance of downregulation of CH25H in DCs for tumor immune evasion and resistance to therapy.

Angiotensin II signaling up-regulates the immediate early transcription factor ATF3 in the left but not the right atrium.

The atria respond to various pathological stimuli including pressure and volume overload with remodeling and dilatation. Dilatation of the left atrium is associated with atrial fibrillation. The mechanisms involved in chamber-specific hypertrophy are largely unknown. Angiotensin II is hypothesized to take part in mediating this response. ATF3 is an immediate early gene found at the receiving end of multiple stress and growth stimuli. Here we characterize ATF3 as a direct target gene for angiotensin II. ATF3 expression is regulated by angiotensin receptor-mediated signaling in vivo and in vitro at the transcriptional level. ATF3 induction is mediated by cooperation between both the AT(1A) and AT2 receptor subtypes. While AT2R blocker (PD123319) efficiently blocks ATF3 induction in response to angiotensin II injection, it results in an increase in blood pressure indicating that the effect of angiotensin II on ATF3 is independent of its effect on blood pressure. In contrast to adrenergic stimulation that induces ATF3 in all heart chambers, ATF3 induction in response to angiotensin II occurs primarily in the left chambers. We hypothesize that the activation of differential signaling pathways accounts for the chamber-specific induction of ATF3 expression in response to angiotensin II stimulation. Angiotensin II injection rapidly activates the EGFR-dependent pathways including ERK and PI3K-AKT in the left but not the right atrium. EGF receptor inhibitor (Gefitinib/Iressa) as well as the AKT inhibitor (Triciribine) significantly abrogates ATF3 induction by angiotensin II in the left chambers. Collectively, our data strongly place ATF3 as a unique nuclear protein target in response to angiotensin II stimulation in the atria. The spatial expression of ATF3 may add to the understanding of the signaling pathways involved in cardiac response to neuro-hormonal stimulation, and in particular to the understanding of left atrial-generated pathology such as atrial fibrillation.

Activating transcription factor 3 confers protection against ventilator-induced lung injury.

RATIONALE: Ventilator-induced lung injury (VILI) significantly contributes to mortality in patients with acute respiratory distress syndrome, the most severe form of acute lung injury. Understanding the molecular basis for response to cyclic stretch (CS) and its derangement during high-volume ventilation is of high priority. OBJECTIVES: To identify specific molecular regulators involved in the development of VILI. METHODS: We undertook a comparative examination of cis-regulatory sequences involved in the coordinated expression of CS-responsive genes using microarray analysis. Analysis of stretched versus nonstretched cells identified significant enrichment for genes containing putative binding sites for the transcription factor activating transcription factor 3 (ATF3). To determine the role of ATF3 in vivo, we compared the response of ATF3 gene-deficient mice to wild-type mice in an in vivo model of VILI. MEASUREMENTS AND MAIN RESULTS: ATF3 protein expression and nuclear translocation is increased in the lung after mechanical ventilation in wild-type mice. ATF3-deficient mice have greater sensitivity to mechanical ventilation alone or in conjunction with inhaled endotoxin, as demonstrated by increased cell infiltration and proinflammatory cytokines in the lung and bronchoalveolar lavage, and increased pulmonary edema and indices of tissue injury. The expression of stretch-responsive genes containing putative ATF3 cis-regulatory regions was significantly altered in ATF3-deficient mice. CONCLUSIONS: ATF3 deficiency confers increased sensitivity to mechanical ventilation alone or in combination with inhaled endotoxin. We propose ATF3 acts to counterbalance CS and high volume-induced inflammation, dampening its ability to cause injury and consequently protecting animals from injurious CS.

ATF3, a hub of the cellular adaptive-response network, in the pathogenesis of diseases: is modulation of inflammation a unifying component?

Activating transcription factor 3 (ATF3) gene encodes a member of the ATF family of transcription factors and is induced by various stress signals. All members of this family share the basic region-leucine zipper (bZip) DNA binding motif and bind to the consensus sequence TGACGTCA in vitro. Previous reviews and an Internet source have covered the following topics: the nomenclature of ATF proteins, the history of their discovery, the potential interplays between ATFs and other bZip proteins, ATF3-interacting proteins, ATF3 target genes, and the emerging roles of ATF3 in cancer and immunity (see footnote 1). In this review, we present evidence and clues that prompted us to put forth the idea that ATF3 functions as a "hub" of the cellular adaptive-response network. We will then focus on the roles of ATF3 in modulating inflammatory response. Inflammation is increasingly recognized to play an important role for the development of many diseases. Putting this in the context of the hub idea, we propose that modulation of inflammation by ATF3 is a unifying theme for the potential involvement of ATF3 in various diseases.