ABSTRACT
Special AT-rich sequence-binding protein 2 (Satb2) is a transcriptional regulator and people with SATB2 mutation or duplication could display epilepsy. However, whether Satb2 is related with epilepsy and its mechanisms are largely unexplored. Here we found that the expression of Satb2 was decreased following the neuronal hyperactivities. Ablation of Satb2 in mice would decrease incidence and stage of seizure induced by intraperitoneal injection of pentylenetetrazol (PTZ). At cellular levels, we found pyramidal neuronal excitability and excitatory synaptic inputs in CA1 were decreased in Satb2 mutant mice. Taking together, we proved that deletion of Satb2 in mice increased PTZ seizure threshold probably by modulating neuronal excitability.
Subject(s)
Matrix Attachment Region Binding Proteins/drug effects , Pentylenetetrazole/pharmacology , Pyramidal Cells/metabolism , Seizures/metabolism , Transcription Factors/drug effects , Animals , Epilepsy/metabolism , Hippocampus/drug effects , Hippocampus/metabolism , Matrix Attachment Region Binding Proteins/metabolism , Mice, Transgenic , Pyramidal Cells/drug effects , Seizures/chemically induced , Transcription Factors/metabolismABSTRACT
"Triple-negative" (TN) breast cancers, which are characterized by estrogen receptor (-), progesterone receptor (-), and human epidermal growth factor receptor 2 (-), are typically associated with poor prognosis because of their aggressive tumor phenotypes. In recent years, the number of patients with breast cancers has remarkably increased, but there are only few available drugs for treatment of TN breast cancers. The development of novel drugs targeting TN breast cancer is urgently required. In the present study, we focused on the function of special AT-rich sequence binding protein 1 (SATB1) as a target molecule for the treatment of TN breast cancers. By recruiting chromatin remodeling enzymes and transcriptional factors, SATB1 regulates the expression of >1,000 genes related to cell growth and translocation. We synthesized a decoy DNA against SATB1, including the recognition sequence of SATB1. We examined the inhibitory effects of the decoy DNAs on cellular proliferation of a TN metastatic breast cancer cell line (MDA-MB-231). SATB1-decoy DNA inhibited the proliferation of MDA-MB-231 cells. Especially, it was significant that SATB1-decoy DNA drastically reduced the invasive and metastatic capacity of MBA-MB-231 cells. Further, in the case of MCF7 cells (SATB1-negative breast cancer cell line), SATB1-decoy DNA did not exhibit any inhibitory effect. These data suggest that SATB1-decoy DNA may be an effective candidate for use as a molecular-targeting drug for treatment of TN breast cancer.
Subject(s)
Breast Neoplasms/pathology , Breast Neoplasms/therapy , Genetic Therapy , Matrix Attachment Region Binding Proteins/antagonists & inhibitors , Matrix Attachment Region Binding Proteins/metabolism , Oligonucleotides/pharmacology , Breast Neoplasms/genetics , Cell Line, Tumor , Cell Proliferation/drug effects , Connective Tissue Growth Factor/drug effects , Connective Tissue Growth Factor/metabolism , DNA/chemical synthesis , DNA/genetics , DNA/pharmacology , DNA-Binding Proteins/antagonists & inhibitors , DNA-Binding Proteins/drug effects , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Down-Regulation/drug effects , Female , Humans , Matrix Attachment Region Binding Proteins/drug effects , Matrix Attachment Region Binding Proteins/genetics , Neoplasm Invasiveness/genetics , Oligonucleotides/chemical synthesis , Oligonucleotides/genetics , Receptor, ErbB-2/drug effects , Receptor, ErbB-2/metabolism , S100 Calcium-Binding Protein A4 , S100 Proteins/drug effects , S100 Proteins/metabolism , Vascular Endothelial Growth Factor B/drug effects , Vascular Endothelial Growth Factor B/metabolismABSTRACT
Activity of the estrogen receptor (ER) is regulated through interaction with coactivators and corepressors. These proteins are present in large complexes, suggesting functional interactions among various cofactors. Scaffold attachment factors B1 and B2 (SAFB1/2) and nuclear receptor corepressor (N-CoR) function as ERalpha corepressors--they directly interact with ERalpha, and repress transcription via repression domains. We asked the question whether SAFB1/2 and N-CoR could directly interact with each other, and whether this interaction results in altered repressive activities. Employing coimmunoprecipitation, cofractionation, and colocalization experiments, we have shown that SAFB1/2 interact with the nuclear receptor corepressor N-CoR. This interaction was direct, and was mediated in vitro and in vivo through the C-terminal region of SAFB1 (amino acids 600-915 and the N-terminal region of N-CoR (amino acids 1-373)). Decrease of SAFB1 or N-CoR expression by small interfering RNA resulted in an increase of the estrogen response in reporter assays, confirming prior data that both proteins are attenuating estrogen-mediated induction of genes. Importantly, the effect of SAFB1 on this attenuation was significantly decreased in the presence of N-CoR small interfering RNA. Using chromatin immunoprecipitation assays, we observed that SAFB1/2 and N-CoR were recruited to the pS2 promoter in the absence of estrogen, and this recruitment was enhanced in the presence of Tamoxifen. Detailed kinetic studies showed that the addition of estrogen resulted in the concurrent release of SAFB1/2 and N-CoR from the promoter. Finally, we measured expression of SAFB1/2 and N-CoR in 289 clinical breast cancer specimens, and detected a strong and highly significant correlation between their expression levels. Taken together, our studies demonstrate that SAFB1/2 and N-CoR interact, and that this interaction is, at least in part, necessary for SAFB1's repressive activities. The coexpression of these proteins in breast cancer specimens, and the combined recruitment (and release) of SAFB1/2 and N-CoR furthermore suggests that this interaction has functional relevance.
Subject(s)
Breast Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Matrix Attachment Region Binding Proteins/metabolism , Nuclear Matrix-Associated Proteins/metabolism , Nuclear Proteins/metabolism , Receptors, Estrogen/metabolism , Repressor Proteins/metabolism , Aged , Aged, 80 and over , Antineoplastic Agents, Hormonal/pharmacology , Breast Neoplasms/chemistry , Breast Neoplasms/metabolism , Chromatin Immunoprecipitation , Down-Regulation , Estrogen Receptor alpha/metabolism , Estrogens/pharmacology , Female , Humans , Matrix Attachment Region Binding Proteins/analysis , Matrix Attachment Region Binding Proteins/drug effects , Middle Aged , Nuclear Matrix-Associated Proteins/analysis , Nuclear Matrix-Associated Proteins/drug effects , Nuclear Proteins/analysis , Nuclear Proteins/genetics , Nuclear Receptor Co-Repressor 1 , Promoter Regions, Genetic/drug effects , RNA, Small Interfering/genetics , RNA, Small Interfering/pharmacology , Receptors, Estrogen/analysis , Receptors, Estrogen/drug effects , Repressor Proteins/analysis , Repressor Proteins/genetics , Tamoxifen/pharmacologyABSTRACT
3-Aminobenzamide (3AB) is an inhibitor of poly (ADP-ribose) polymerase (PARP), an enzyme implicated in the maintenance of genomic integrity, which is activated in response to radiation-induced DNA strand breaks. cDNA macroarray membranes containing 1536 clones were used to characterize the gene expression profiles displayed by mouse BALB/3T3 fibroblasts (A31 cell line) in response to ionizing irradiation alone or in combination with 3AB. A31 cells in exponential growth were pre-treated with 3AB 4mM 1h before gamma-irradiation (4Gy), remaining in culture during 6h until harvesting time. A31 cells treated with 3AB alone presented a down-regulation in genes involved in protein processing and cell cycle control, while an up-regulation of genes involved in apoptosis and related to DNA/RNA synthesis and repair was verified. A31 cells irradiated with 4Gy displayed 41 genes differentially expressed, being detected a down-regulation of genes involved in protein processing and apoptosis, and genes controlling the cell cycle. Concomitantly, another set of genes for protein processing and related to DNA/RNA synthesis and repair were found to be up-regulated. A positive or negative interaction effect between 3AB and radiation was verified for 29 known genes. While the combined treatment induced a synergistic effect on the expression of LCK proto-oncogene and several genes related to protein synthesis/processing, a negative interaction effect was found for the expression of genes related to cytoskeleton and extracellular matrix assembly (SATB1 and Anexin III), cell cycle control (tyrosine kinase), and genes participating in DNA/RNA synthesis and repair (RNA helicase, FLAP endonuclease-1, DNA-3 glycosylase methyladenine, splicing factor SC35 and Soh1). The present data open the possibility to investigate the direct participation of specific genes, or gene products acting in concert in the mechanism underlying the cell response to radiation-induced DNA damage under the influence of PARP inhibitor.
