ABSTRACT
Objective To determine the location of PQ-LRP protein in Microsporum canis using the enhanced green fluorescent protein(EGFP)as a marker.Methods The total RNA was extracted from Microsporum canis,and reversely transcribed into cDNA.The PQ-LRP gene was amplified by PCR using the above cDNA as the template.The fusion gene of PQ-LRP gene and EGFP gene was linked to the plasmid pCAMBIA 1300.Microsporum canis was subjected to Agrobacterium tumefaciens-mediated transformation,in order to achieve the integrated expression of the fusion gene LRP-EGFP in Microsporum canis under the regulation by the fungal universal promoter Ptrpc and terminator Ttrpc.Laser-scanning confocal microscopy was conducted to determine the cellular localization of the fusion protein.Results The expression vector pCAMBIA-LRP-EGFP was successfully constructed,and the fusion gene LRP-EGFP was expressed integratedly in Microsporum canis.Laser-scanning confocal microscopy showed that fluorescence signals of LRP-EGFP were concentrated on the cell membrane of Microsporum canis,giving a granular or cluster-like appearance.Conclusion The infusion gene LRP-EGFP can be successfully expressed in Microsporum canis,and PQ-LRP protein is located on the cell membrane of Microsporum canis.
ABSTRACT
AT-motif binding factor 1 (ATBF1), also named as zinc finger homeobox 3 (ZFHX3), is reported as a transcriptional regulator with the highest molecular weight. ATBF1 regulates mRNA transcription by binding to AT-rich motif of gene promoter. In recent years, more and more studies have shown that ATBF 1 gene is an important candidate tumor suppressor gene. Frequent mutations and loss of heterozygosity (LOH) are found in ATBF 1 gene in multiple tumors, including prostate cancer, gastric cancer, breast cancer, cervical cancer and colorectal cancer. The gene expression and protein localization of ATBF1 are associated with the clinical diagnosis and treatment of tumors. Moreover, ATBF1 plays an essential role in multiple signaling pathways to regulate cell proliferation, differentiation and apoptosis. This review summarizes the research progress of ATBF 1 gene in malignant tumors from four aspects: DNA, mRNA, protein and molecular mechanism, in order to comprehesively understand the anti-cancer mechanism of ATBF1 and its clinical significance in diagnosis and treatment of tumors.
ABSTRACT
PURPOSE: Establishment of the human liver cell lines which permanently express the HCV proteins is important for the large scale production of viral antigen and analysis of the mechanism of hepatocellular carcinogenesis by HCV. Here, we attempted to establish the human hepatoblastoma cell lines which stably express the HCV core protein and examined the intracellular localization of the core protein. MATERIALS AND METHODS: The cDNA of HCV core protein and neomycin resistance gene were expressed in HepG2 cells by the SRalpha promoter and human EF-1alpha gene promoter, respectively. The core protein was detected by immunofluorescence assay and western blotting. RESULTS: We obtained several HepG2 cell clones which express HCV core protein stably. In transient expression assay, the core protein was localized in the cytoplasm in about 90%, and localized in the nucleus in about 10% of the core-expressing HepG2 cells. But, in the stably expressing HepG2 cell clones, the core protein was localized only in the cytoplasm. No HepG2 cell containing core protein in the nucleus was found in all of the cells which stably express the core protein. CONCLUSION: The EF-1alpha gene promoter is highly efficient in the colony formation by neomycin resistance gene and is very useful for the isolation of human liver cell clones which express foreign genes stably. HCV core protein is localized in both nuclear and cytoplasm of human liver cell in short term but the cells containing the core protein in nucleus seem to disappear in long term culture.