ABSTRACT
<p><b>OBJECTIVE</b>To identify the expression of Drosophila Eyes Absent Homologue 2 (EYA2) in non-small cell lung cancer (NSCLC) and to investigate its correlation with clinical parameters.</p><p><b>METHODS</b>59 fresh specimens of lung cancer and paired normal lung tissue were obtained from 59 NSCLC cases treated in the department of thoracic surgery in our hospital from June 2006 to October 2007. Western blotting and immunohistochemistry were used to assay the specimens with goat anti-human EYA2 polyclone antibody. Clinicopathological parameters were collected and the correlation with EYA2 expression was subsequently analyzed.</p><p><b>RESULTS</b>The expression of EYA2 was detected in cytoplasm and nucleus of the cancer cells, but mostly in cytoplasm. Western blotting and immunohistochemistry showed the expression of EYA2 in NSCLC was increased and correlated with pathological type, but not with gender, age, pTNM stage, histological differentiation and lymph node metastasis. EYA2 expression was significantly up-regulated in adenocarcinoma, while not changed in lung squamous cell carcinoma.</p><p><b>CONCLUSION</b>The results of this study suggest that expression of EYA2 in lung adenocarcinoma is augmented. EYA2 is likely participating in the development of lung adenocarcinoma as a transcriptional activator.</p>
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Adenocarcinoma , Metabolism , Pathology , Carcinoma, Non-Small-Cell Lung , Metabolism , Pathology , Carcinoma, Squamous Cell , Metabolism , Pathology , Cytoplasm , Metabolism , Intracellular Signaling Peptides and Proteins , Metabolism , Lung , Metabolism , Lung Neoplasms , Metabolism , Pathology , Lymphatic Metastasis , Neoplasm Staging , Nuclear Proteins , Metabolism , Protein Tyrosine Phosphatases , Metabolism , Up-RegulationABSTRACT
<p><b>OBJECTIVES</b>To study the effects of trichostatin A (TSA) on protein-protein interaction between HBx and histone deacetylase protein 1 (HDAC1).</p><p><b>METHODS</b>Both HBx and HDAC1 expressing vectors were constructed by the method of routine molecular cloning. The expression of HBx and HDAC1 were observed by Western blot assay. The protein-protein interaction was tested between HBx and HDAC1 by GST pull-down in vitro as well as co-immunoprecipitation in vivo.</p><p><b>RESULTS</b>Both HBx and HDAC1 expressing vectors were successfully constructed. Protein-protein interaction between HBx and HDAC1 existed both in vitro and in vivo. TSA, an inhibitor of HDAC1, had no effect on the interaction between HBx and HDAC1.</p><p><b>CONCLUSIONS</b>HBx interacts with HDAC1 in vivo and in vitro in a non- TSA dependent way.</p>
Subject(s)
Humans , Histone Deacetylase 1 , Metabolism , Hydroxamic Acids , Metabolism , Immunoprecipitation , Plasmids , Protein Interaction Mapping , Trans-Activators , MetabolismABSTRACT
The complete coding sequences of Eya gene family was amplified by standard PCR fromhuman tissues or cells cDNA library.The product of PCR was cloned into the eukaryotic expression vector pcDNA3-FLAG,generating pcDNA3-FLAG-Eya1～4.Thenhuman embryo kidney 293T cells were transfected with the recombinant plasmids and the expression of Eya genes were identified by Western blot.Transcriptional assay using a reporter containing myogenin enhance factor indicated that expression of Eya cooperation with Six in 293T cells affected the Myogenin gene expression.The expression vectors of Eya genes were constructed and confirmed by restriction enzyme digestion and DNA sequence analysis.Transcriptional assay using a reporter containing myogenin enhance factor indicated that expression of Eya in coordination with Six in 293T cells stimulated the Myogenin gene expression.Eya proteins are transcriptional activator of Six and can improve the activity of myogenin promoter.
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the features of HBx protein distributed in liver cells and its expression in E. coli.</p><p><b>METHODS</b>The expression vectors encoding the full length HBx and its mutants were constructed by the routine molecular cloning method. HBx protein expression was detected using Western blotting. The distribution feature of HBx protein in liver cells was examined using the fluorescence confocal microscopy. A series of purified HBx fusion proteins were obtained by glutathione-sepharose 4B affinity chromatography.</p><p><b>RESULTS</b>The expression vectors were successfully constructed for the full length HBx and its mutants. HBx was found distributed uniformly in the nuclei but granularly in the cytoplasm of the liver cells. Under optimal conditions, the mutant GST-HBx (72-120aa) was easily degraded.</p><p><b>CONCLUSION</b>This study may provide a basis for further study on the biological function of HBx at the protein level.</p>
Subject(s)
Humans , Carcinoma, Hepatocellular , Pathology , Cell Line , Cloning, Molecular , Escherichia coli , Metabolism , Genetic Vectors , Glutathione Transferase , Genetics , Hepatocytes , Cell Biology , Metabolism , Liver , Cell Biology , Liver Neoplasms , Pathology , Mutation , Recombinant Fusion Proteins , Genetics , Trans-Activators , Genetics , Tumor Cells, CulturedABSTRACT
Estrogen receptor alpha (ERalpha) has been a primary target of treatment as well as a prognostic indicator for breast cancer. The level of human X-box binding protein 1 (XBP-1) mRNA was related with that of ERalpha in breast tumors and was over-expressed in some breast tumors. These previous studies suggested that XBP-1 may interact with ERalpha. XBP-1 has two isoforms, XBP-1S and XBP-1U, as the result of unique splicing. GST pull-down assay showed that both XBP-1S and XBP-1U bound to ERalpha in vitro. The binding of XBP-1S to ERalpha was stronger than that of XBP-1U to ERalpha. Co-immunoprecipitation revealed that the binding was in a ligand-independent manner. XBP-1S and XBP-1U interacted with the region of ERalpha that contains a DNA-binding domain. The ERalpha-interacting regions on XBP-1S and XBP-1U have been mapped to two regions, the N-terminal basic region leucine zipper domain (bzip) and the C-terminal activation domain. These findings suggest that XBP-1S and XBP-1U may participate in ERalpha signaling pathway through the mediation of ERalpha.
Subject(s)
Female , Humans , Breast Neoplasms , Genetics , Metabolism , Cell Line, Tumor , DNA-Binding Proteins , Genetics , Metabolism , Estrogen Receptor alpha , Genetics , Metabolism , Protein Interaction Domains and Motifs , Physiology , RNA, Messenger , Genetics , Regulatory Factor X Transcription Factors , Signal Transduction , Transcription Factors , Genetics , Metabolism , X-Box Binding Protein 1ABSTRACT
<p><b>OBJECTIVE</b>To observe the changes in peripheral blood T lymphocyte rDNA transcription activity and to study the significance of immune monitoring for patients with chronic benzene poisoning.</p><p><b>METHODS</b>Venous blood samples were withdrawn from 39 patients with chronic benzene poisoning, 20 patients with malignant disease and 22 healthy controls. Cell culture, argyrophil staining method, I-CLQ cell image analysis system were used in this study. rDNA transcription activity which was expressed by the ratio of integrated area (IA) of nucleolus to that of nucleus, and the ratio of integrated optical density (IOD) of argyrophilic nucleolus to that of argyrophilic nucleus.</p><p><b>RESULTS</b>(1) The value of IA and IOD in chronic benzene poisoning patients (7.95% +/- 1.13% and 7.15% +/- 1.15% respectively) were lower than those in controls (9.59% +/- 1.26% and 8.92% +/- 1.18% respectively), P < 0.01. The value of IA and IOD in chronic moderate benzene poisoning patients (6.54% +/- 0.88%) and (5.47% +/- 0.80%) were lower than those to be observed (7.98% +/- 1.06% and 7.13% +/- 0.96% respectively) as well as in mild poisoning patients (8.19% +/- 1.06% and 7.44% +/- 1.06% respectively), P < 0.05. (2) The value of IA and IOD in malignant group (4.10% +/- 1.50%, 3.67% +/- 1.42%) were much more lower than those in controls and in chronic benzene poisoning patients (P < 0.01).</p><p><b>CONCLUSION</b>rDNA transcription activity may be an index to monitor the cellular immune function of chronic benzene poisoning patients.</p>