关键词： Antagonists   Cancer   Electrostatics   Hydrophobicity   Peptides and proteins  

摘要： Activating transcription factor 3 (ATF3) is an activation transcription factor/cyclic adenosine monophosphate (cAMP) responsive element-binding (CREB) protein family member. It is recognized as an important regulator of cancer progression by repressing expression of key inflammatory factors such as interferon-gamma and chemokine (C-C motif) ligand 4 (CCL4). Here, we describe a novel library screening approach that probes individual leucine zipper components before combining them to search exponentially larger sequence spaces not normally accessible to intracellular screening. To do so, we employ two individual semirational library design approaches and screen using a protein-fragment complementation assay (PCA). First, a 248,832-member library explored 12 amino acid positions at all five a positions to identify those that provided improved binding, with all e/g positions fixed as Q, placing selection pressure onto the library options provided. Next, a 59,049-member library probed all ten e/g positions with 3 options. Similarly, during e/g library screening, a positions were locked into a generically bindable sequence pattern (AIAIA), weakly favoring leucine zipper formation, while placing selection pressure onto e/g options provided. The combined a/e/g library represents similar to 14.7 billion members, with the resulting peptide, ATF3W_aeg, binding ATF3 with high affinity (T (m) = 60 C-degrees;K (d) = 151 nM) while strongly disfavoring homodimerization. Moreover, ATF3W_aeg is notably improved over component PCA hits, with target specificity found to be driven predominantly by electrostatic interactions. The combined a/e/g exponential library screening approach provides a robust, accelerated platform for exploring larger peptide libraries, toward derivation of potent yet selective antagonists that avoid homoassociation to provide new insight into rational peptide design.

关键词： HVPRF   Neuropathic pain   Autophagy   ATF3   TNF-alpha   IL-10  

摘要： Neuropathic pain (NeP) is intractable for which many therapies are ineffective. High-voltage pulsed radiofrequency (HVPRF) on dorsal root ganglion (DRG) is considered an effective treatment for NeP. The aim of this study is to explore the therapeutic voltage for the optimal efficacy of PRF and the underlying mechanisms. The radiofrequency electrode was placed close to the L5 DRG of rats with spared nerve injury (SNI) and emitted current by the corresponding voltage in different groups. Four different voltages (45 V, 65 V, 85 V, and 100 V) of PRF on DRG significantly alleviated the SNI-induced NeP, reduced the levels of activating transcription factor 3 (ATF3) in DRG, improved the ultrastructure of DRG, and promoted autophagy in spinal microglia to varying degrees and partially reversed the increased expression of TNF-alpha and the reduced expression of IL-10 in spinal cord dorsal horn (SCDH). The beneficial effect of 85V-PRF was superior to those of other three PRF treatments. The underlying mechanisms may be related to repairing the DRG damage and improving the DRG ultrastructure while regulating spinal microglial autophagy and thereby alleviating neuroinflammation.

关键词： ATF3   ILF3   NEAT1   Sepsis   Macrophage polarization  

摘要： BackgroundSepsis is a systemic inflammatory response which is frequently associated with acute lung injury (ALI). Activating transcription factor 3 (ATF3) promotes M2 polarization, however, the biological effects of ATF3 on macrophage polarization in sepsis remain ***-stimulated macrophages and a mouse model of cecal ligation and puncture (CLP)-induced sepsis were generated as in vitro and in vivo models, respectively. qRT-PCR and western blot were used to detect the expression of ATF3, ILF3, NEAT1 and other markers. The phenotypes of macrophages were monitored by flow cytometry, and cytokine secretion was measured by ELISA assay. The association between ILF3 and NEAT1 was validated by RIP and RNA pull-down assays. RNA stability assay was employed to assess NEAT1 stability. Bioinformatic analysis, luciferase reporter and ChIP assays were used to study the interaction between ATF3 and ILF3 promoter. Histological changes of lung tissues were assessed by H&E and IHC analysis. Apoptosis in lungs was monitored by TUNEL ***3 was downregulated, but ILF3 and NEAT1 were upregulated in PBMCs of septic patients, as well as in LPS-stimulated RAW264.7 cells. Overexpression of ATF3 or silencing of ILF3 promoted M2 polarization of RAW264.7 cells via regulating NEAT1. Mechanistically, ILF3 was required for the stabilization of NEAT1 through direct interaction, and ATF3 was a transcriptional repressor of ILF3. ATF3 facilitated M2 polarization in LPS-stimulated macrophages and CLP-induced septic lung injury via ILF3/NEAT1 ***3 triggers M2 macrophage polarization to protect against the inflammatory injury of sepsis through ILF3/NEAT1 axis.

关键词： TBBPA   Mitophagy deficiency   Neuronal cytotoxicity   PINK1/Parkin   ATF3  

摘要： Tetrabromobisphenol A (TBBPA), as a widely used brominated flame retardant, has been implicated as a potential neurotoxicant. However, the mechanism of TBBPA-induced neurotoxicity has not been fully elucidated yet. In this study, using mouse hippocampal neuron cell HT22 as the in vitro model, the neuronal cytotoxicity of TBBPA and the mechanism by focusing on mitophagy have been studied. We found that neuronal cytotoxic effects were indeed induced by TBBPA exposure at concentrations of >20 IrM for 24 h, including decreased cell viability (to 92.38 % at 20 IrM;18.25 % at 80 IrM), enhanced ROS (enhanced 53.26 % at IC50 of 60 IrM, compared with that in the control group) and mitochondrial ROS (mtROS) levels (enhanced 24.12 % at 60 IrM), reduced mitochondrial membrane potential (MMP) (decreased 33.60 % at 60 IrM). As a protective mechanism in cells, autophagy was initiated;however, mitophagy was inhibited, where PINK1 (PINK1-Parkin activation is critical in the depolarized MMP-induced mitophagy) expression was found to be repressed and decreased, further leading to the failure of Parkin recruitment to the damaged mitochondria. Mitophagy activator, nicotinamide mono nucleotide (13-NMN) that activates the PINK1-Parkin pathway, could alleviate TBBPA-induced mitophagy deficiency and further reduce the neuronal cytotoxicity, demonstrating that TBBPA-induced PINK1-Parkin-mediated mitophagy deficiency contributed to the neuronal cytotoxicity. Furthermore, we found TBBPA caused the upregulation of Atf3 (activating transcription factor 3) gene transcription and expression levels, alongside reduced Pink1 levels;whereas enhanced Pink1 transcript levels were observed after ATF3 depletion even under TBBPA treatment, demonstrating TBBPA-induced overexpression of ATF3 should be responsible for the reduced PINK1 expression. Therefore, for the first time, here we demonstrate that TBBPA can inhibit PINK1-Parkinmediated mitophagy via upregulating ATF3 expression, which further contributes

关键词： ATF3   IGF2BP2   acute lung injury   circEXOC5   ferroptosis   sepsis  

摘要： Background Patients with sepsis resulting in acute lung injury (ALI) usually have increased mortality. Ferroptosis is a vital regulator in sepsis-induced ALI. Exploring the association of ferroptosis and sepsis-induced ALI is crucial for the management of sepsis-induced *** Whole blood was collected from sepsis patients. Mice were treated with cecal ligation and puncture (CLP) to model sepsis. Primary murine pulmonary microvascular endothelial cells were treated with lipopolysaccharide as a cell model. Ferroptosis was evaluated by analyzing levels of iron, malonaldehyde, glutathione, nonheme iron, ferroportin, ferritin, and GPX4. Hematoxylin and eosin and Masson's trichrome staining were applied to examine lung injury and collagen deposition. Cell apoptosis was analyzed by caspase-3 activity and TUNEL assays. Gene regulatory relationship was verified using RNA pull-down and immunoprecipitation *** CircEXOC5 was highly expressed in sepsis patients and CLP-treated mice, in which knockdown alleviated CLP-induced pulmonary inflammation and injury, and ferroptosis. CircEXOC5 recruited IGF2BP2 to degrade ATF3 mRNA. The demethylase ALKBH5 was responsible for circEXOC5 upregulation through demethylation. CircEXOC5 silencing significantly improved sepsis-induced ALI and survival rate of mice by downregulating *** ALKBH5-mediated upregulation of circEXOC5 exacerbates sepsis-induced ALI by facilitating ferroptosis through IGF2BP2 recruitment to degrade ATF3 mRNA. Sepsis primarily causes acute lung injury (ALI). Here, we demonstrate that circEXOC5 promotes ferroptosis through the IGF2BP2/ATF3 axis to aggravate sepsis-induced ALI. Our findings provide novel mechanistic insights into sepsis-induced ALI and suggest potential therapeutic targets.

关键词： Cancer   Cells   Peptides and proteins   Rodent models   Tumors  

摘要： Homo sapiens caseinolytic protease P (HsClpP) activation is a promising strategy for colon cancer treatment. In this study, CCG1423 was identified as a selective activator of HsClpP. After optimization, NCA029 emerged as the most potent compound, with an EC50 of 0.2 mu M against HsClpP. Molecular dynamics revealed that the affinity of NCA029 for the YYW aromatic network is crucial for its selectivity toward HsClpP. Furthermore, NCA029 displayed favorable pharmacokinetics and safety profiles and significantly inhibited tumor growth in HCT116 xenografts, resulting in 83.6% tumor inhibition. Mechanistically, NCA029 targeted HsClpP, inducing mitochondrial dysfunction and activating the ATF3-dependent integrated stress response, ultimately causing cell death in colorectal adenocarcinoma. These findings highlight NCA029 as an effective HsClpP activator with potential for colon cancer therapy.

关键词： Breast cancer   Bone metastasis   miR-4638-3p   ATF3   Caudal artery injection  

摘要： PurposeSkeletal metastases are increasingly reported in metastatic triple-negative breast cancer (BC) patients. We previously reported that TGF-beta 1 sustains activating transcription factor 3(ATF3) expression and is required for cell proliferation, invasion, and bone metastasis genes. Increasing studies suggest the critical regulatory function of microRNAs (miRNAs) in governing BC pathogenesis. TGF-beta 1 downregulated the expression of miR-4638-3p, which targets ATF3 in human BC cells (MDA-MB-231). In the present study, we aimed to identify the functional role of miR-4638-3p in BC bone metastasis by the caudal artery injection of the MDA-MB-231 cells overexpressing mir-4638 in the ***-MB-231 cells overexpressing miR-4638 were prepared by stable transfections. Reverse transcriptase quantitative PCR was carried out to determine the expression of endogenous miR-4638-3p and bone resorption marker genes. X-ray, micro-CT, and Hematoxylin & Eosin studies were used to determine osteolytic lesions, trabecular structure, bone mineral density, and micrometastasis of *** mice injected with MDA-MB-231 cells overexpressing miR-4638-3p decreased the expression of bone resorption marker genes, compared to MDA-MB-231 cells injection. Reduced osteolytic lesions and restored bone density by MDA-MB-231 cells overexpressing miR-4638-3p were observed. Similarly, the mice injected with MDA-MB-231 cells overexpressing miR-4638-3p showed a better microarchitecture of the trabecular network. A few abnormal cells seen in the femur of MDA-MB-231 cells-injected mice were not found in MDA-MB-231 cells overexpressing *** identified functional role of ATF3 targeting miR-4638-3p in BC bone metastasis in vivo suggests its candidature as BC therapeutics in the future.

关键词： Intestinal ischemia reperfusion injury   Apigenin-7-O-glucoside   Nanoparticle   Ferroptosis   Oxidative stress  

摘要： Intestinal ischemia reperfusion injury (II/R injury) is a common and intractable pathophysiological process in critical patients, for which exploring new treatments and mechanisms is of great importance to improve treatment outcomes. Apigenin-7-O-Glucoside (AGL) is a sugar derivative of apigenin natural product with various pharmacological activities to protect against intestinal diseases. In this study, we synthesized two amphiphilic molecules, namely DTPA-N10-10 and mPEG-TK-DA, which can scavenge free radicals and reactive oxygen species (ROS). They were successfully encapsulated AGL through self-assembly, resulting in the formation of multi-site ROS scavenging nanoparticles called PDN@AGL. In vitro and in vivo experiments demonstrated that PDN@AGL could protect intestinal tissues by reducing lipid peroxidation, lowering ROS levels and inhibiting ferroptosis during II/R injury. Furthermore, our study revealed, for the first time, that the regulation of the ATF3/SLC7A11 pathway by PDN@AGL may play a crucial role in mitigating II/R injury. In conclusion, our study confirmed the beneficial effects of PDN@AGL in combating II/R injury through the ATF3/SLC7A11-mediated regulation of ferroptosis and oxidative stress. These findings lay the groundwork for the potential application of PDN@AGL in the treatment of II/R injury.

关键词： Ferroptosis   SIRT1   AROS   ATF3   Glioma   Oxidative stress   NAD plus  

摘要： Ferroptosis is a type of programmed cell death resulting from iron overload-dependent lipid peroxidation, and could be promoted by activating transcription factor 3 (ATF3). SIRT1 is an enzyme accounting for removing acetylated lysine residues from target proteins by consuming NAD+, but its role remains elusive in ferroptosis and activating ATF3. In this study, we found SIRT1 was activated during the process of RSL3-induced glioma cell ferroptosis. Moreover, the glioma cell death was aggravated by SIRT1 activator SRT2183, but suppressed by SIRT inhibitor EX527 or when SIRT1 was silenced with siRNA. These indicated SIRT1 sensitized glioma cells to ferroptosis. Furthermore, we found SIRT1 promoted RSL3-induced expressional upregulation and nuclear translocation of ATF3. Silence of ATF3 with siRNA attenuated RSL3-induced increases of ferrous iron and lipid peroxidation, downregulation of SLC7A11 and GPX4 and depletion of cysteine and GSH. Thus, SIRT1 promoted glioma cell ferroptosis by inducting ATF3 activation. Mechanistically, ATF3 activation was reinforced when RSL3-induced decline of NAD+ was aggravated by FK866 that could inhibit NAD + synthesis via salvage pathway, but suppressed when intracellular NAD+ was maintained at higher level by supplement of exogenous NAD+. Notably, the NAD + decline caused by RSL3 was enhanced when SIRT1 was further activated by SRT2183, but attenuated when SIRT1 activation was inhibited by EX527. These indicated SIRT1 promoted ATF3 activation via consumption of NAD+. Finally, we found RSL3 activated SIRT1 by inducing reactive oxygen species-dependent upregulation of AROS. Together, our study revealed SIRT1 activated by AROS sensitizes glioma cells to ferroptosis via activation of ATF3-dependent inhibition of SLC7A11 and GPX4.

关键词： ATF3   butyrate   erastin   ferroptosis  

摘要： Ferroptosis is a novel form of programmed cell death triggered by iron-dependent lipid peroxidation and has been associated with various diseases, including cancer. Erastin, an inhibitor of system Xc-, which plays a critical role in regulating ferroptosis, has been identified as an inducer of ferroptosis in cancer cells. In this study, we investigated the impact of butyrate, a short-chain fatty acid produced by gut microbiota, on erastin-induced ferroptosis in lung cancer cells. Our results demonstrated that butyrate significantly enhanced erastin-induced ferroptosis in lung cancer cells, as evidenced by increased lipid peroxidation and reduced expression of glutathione peroxidase 4 (GPX4). Mechanistically, we found that butyrate modulated the pathway involving activating transcription factor 3 (ATF3) and solute carrier family 7 member 11 (SLC7A11), leading to enhanced erastin-induced ferroptosis. Furthermore, partial reversal of the effect of butyrate on ferroptosis was observed upon knockdown of ATF3 or SLC7A11. Collectively, our findings indicate that butyrate enhances erastin-induced ferroptosis in lung cancer cells by modulating the ATF3/SLC7A11 pathway, suggesting its potential as a therapeutic agent for cancer treatment.