DNA-binding Domain Of STAT3 Research Articles

Abstract C173: APE1/Ref-1-STAT3 dual-targeting synergize to effectively inhibit pancreatic cancer cell survival.

Abstract Pancreatic cancer remains a largely incurable disease, with patients facing the worst 5-year survival rate of any cancer. The challenge is to identify the molecular effectors that critically regulate the survival of pancreatic ductal adenocarcinoma (PDAC) cells, to devise effective molecular-targeted strategies that can prevent or minimize the selection of resistant tumor variants, and overcome the protective role of the tumor-associated fibrosis and stroma. Increasing evidence supports the need for strategies targeting multiple molecular effectors in PDAC. Thus, a strategy is to identify critical molecules that regulate multiple signaling mediators (as transcription factors), and on intracellular mechanisms with direct effects on multiple pathways (as antioxidant and redox mediators) critical for PDAC functions. APE1/Ref-1 is a dual function protein, which in addition to DNA repair activity also exerts redox control of transcription factors, including NF- B, p53, AP-1, CREB, HIF-1 and others. Treatment with E3330, a unique small molecule redox signaling inhibitor that recognizes an alternate, redox active conformation of APE1/Ref-1, markedly inhibits the DNA binding and transcriptional activity of NF-κB, AP-1, and HIF-1. Our previous work established APE1/Ref-1 as a potential molecular target in pancreatic cancer. We demonstrated that human adenocarcinoma and peri-pancreatic metastases exhibit increased APE1/Ref-1 expression, and blocking APE1/Ref-1 redox activity delays tumor progression in xenograft models of PDAC, including patient-derived tumor cells. STAT3 is a transcription factor that regulates critical cell functions and has been shown to play important roles in several cancers. STAT3 signaling has been implicated in pancreatic cancer biology, namely by mediating or regulating cell survival, proliferation, tumor angiogenesis and metastasis. Although STAT3 signaling can be engaged and modulated by different processes, the mechanisms regulating STAT3 transcriptional activity in PDAC cells are largely unknown, namely the impact of oxidative stress and its redox status. A recent report demonstrated that STAT3 activity is under redox control and identified the critical oxidation-sensitive cysteines in the STAT3 DNA binding domain. However, the modifier of STAT3 which converts it from an oxidized to a reduced form was not identified. It has been shown that APE1/Ref-1 physically interacts with STAT3 on the VEGF promoter and enhances IL-6-induced DNA binding activity of STAT3 in HepG2 cells. However, it is unknown whether APE-1/Ref-1 is involved in the redox control of STAT3 activity, and whether the cellular redox status affects STAT3 signaling in PDAC cells. Here, we demonstrate that APE1/Ref-1 redox activity regulates the STAT3 DNA binding and transcriptional activity using gene silencing, overexpression of WT or redox-defective APE1/Ref-1, and redox-selective pharmacological inhibition. Blockade of APE1/Ref-1 redox synergizes with STAT3 selective antagonists to markedly inhibit the proliferation and survival of human PDAC cells, inducing cell apoptosis. These studies identify the mechanism by which APE1/Ref-1 regulates STAT3 signaling, and establishes the rationale for the development of APE1/STAT3 dual-targeting strategies for the treatment of pancreatic cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C173.

Dominant-negative mutations in the DNA-binding domain of STAT3 cause hyper-IgE syndrome

Dominant-negative mutations in the DNA-binding domain of STAT3 cause hyper-IgE syndrome

Investigation of latent nuclear localization sequence(NLS) OF STAT3

Objective: To investigate the latent nuclear localization sequence (NLS) OF STAT3. Methods: Clustal X (1.81) was used to alignment the DNA binding domain of the STAT family. According to structure characters, corresponding expression plasmids were constructed via oligonucleotides designed with gene tool software. Through in vitro transfection, images were observed with laser confocal microscopy. Results: homology positions of arginine and lysine were found in DNA binding domain of the stat family. The wild type-STAT3 proteins primarily localized in the cytoplasm and translocated into the nucleus after interleukin-6 stimulation. However, the truncated mutant of DSTAT3-GFP protein was exclusively expressed in the cytoplasm. Conclusion: The potential NLS in the DNA binding domain of STAT3 is exposed to nuclear importing receptor when cells are stinulated by cytokine, which promotes the translocation of STAT3 into nuclear.

The Interaction of Specific Peptide Aptamers With the DNA Binding Domain and the Dimerization Domain of the Transcription Factor Stat3 Inhibits Transactivation and Induces Apoptosis in Tumor Cells

The transcription factor signal transducer and activator of transcription (Stat) 3 is activated through the interleukin-6 family of cytokines and by binding of growth factors to the epidermal growth factor (EGF) receptor. It plays an essential role in embryonic development and assumes specialized tasks in many differentiated tissues. Constitutively activated Stat3 has been found in tumor cell lines and primary tumors and plays a crucial role in tumor cell survival and proliferation. To inhibit the oncogenic action of Stat3 in tumor cells, we have selected short peptides, so-called peptide aptamers, which specifically interact with defined functional domains of this transcription factor. The peptide aptamers were selected from a peptide library of high complexity by an adaptation of the yeast two-hybrid procedure. Peptide aptamers specifically interacting with the Stat3 dimerization domain caused inhibition of DNA binding activity and suppression of transactivation by Stat3 in EGF-responsive cells. Similarly, a peptide aptamer selected for its ability to recognize the Stat3 DNA binding domain inhibited DNA binding and transactivation by Stat3 following EGF stimulation of cells. Peptide aptamers were expressed in bacteria as fusion proteins with a protein transduction domain and introduced into human myeloma cells. This resulted in dose-dependent growth inhibition, down-regulation of Bcl-x(L) expression, and induction of apoptosis. The inhibition of Stat3 functions through the interaction with peptide aptamers counteracts the transformed phenotype and could become useful in targeted tumor therapy.