The DNasel hypersensitive site 2 (HS2) of human β-globin locus control region (LCR) is required fOr the high level expression of human d-globin genes. In the present study, a stage-specific protein factor (LPF-β) wa...The DNasel hypersensitive site 2 (HS2) of human β-globin locus control region (LCR) is required fOr the high level expression of human d-globin genes. In the present study, a stage-specific protein factor (LPF-β) was identified in the nuclear extract prepared from mouse fetal liver at d 18 of gestation, which could bind to the HS2 region of humanβ-globin LCRt We also found that the shift band of LPF-βfactor could be competed by humanβ-globin promoter. However, it couldn’t be competed by human E-globin promoter or by human Aβ-globin promoter. Furthermore, our data demonstrated that the binding-sequence of LPF-d factor is 5’CACACCCTA 3’,which is located at the HS2 region ofβ-LCR (from -10845 to -10853 bp) and humanβ-globin promoter (from -92 to -84 bp). We speculated that these regions containing the CACCC box in both the humallβ-globin promoter and HS2 might function as stage selector elements in the regulation of humanβd-globin switching and the LPF-βfactor might be a stage-specific protein factor involved in the regulation of humanβ-globin gene expression.展开更多
In order to elucidate the molecular mechanisms of globin gene expression during embryonic development, the nuclear extracts from mouse hematopoietic tissue at different stages of development have been prepared. By usi...In order to elucidate the molecular mechanisms of globin gene expression during embryonic development, the nuclear extracts from mouse hematopoietic tissue at different stages of development have been prepared. By using DNase I footprinting and gel mobility shift assays, the binding of protein factors in these extracts to the human βglobin promoter was analyzed. The differences in the binding patterns of protein factors during development were observed. An erythroid-specific and stage-specific nuclear protein in the nuclear extract from d 18 mouse fetal liver was identified, which can bind to the sequence (from -66bp to -90bp) of human β-globin promoter. We therefore speculate that the function of this cis-acting element may be similar to stage selector element (SSE) in chieken βA- promoter.展开更多
Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods The binding of trans-acting factor(s) to two enhancers...Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods The binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer Ⅰ (GPEI) and glutathione S-transferase P enhancer Ⅱ-1 (GPEⅡ-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment. Results Trans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPEⅡ-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver. Conclusion cGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.展开更多
Transfer RNA plays a fundamental role in the protein biosynthesis as an adaptor molecule by functioning as a biological link between the genetic nucleotide sequence in the mRNA and the amino acid sequence in the prote...Transfer RNA plays a fundamental role in the protein biosynthesis as an adaptor molecule by functioning as a biological link between the genetic nucleotide sequence in the mRNA and the amino acid sequence in the protein.To perform its role in protein biosynthesis,it has to be accurately recognized by aminoacyl-tRNA synthetases(aaRSs)to generate aminoacyl-tRNAs(aa-tRNAs).The correct pairing between an amino acid with its cognate tRNA is crucial for translational quality control.Production and utilization of mis-charged tRNAs are usually detrimental for all the species,resulting in cellular dysfunctions.Correct aa-tRNAs formation is collectively controlled by aaRSs with distinct mechanisms and/or other trans-factors.However,in very limited instances,mis-charged tRNAs are intermediate for specific pathways or essential components for the translational machinery.Here,from the point of accuracy in tRNA charging,we review our understanding about the mechanism ensuring correct aa-tRNA generation.In addition,some unique mis-charged tRNA species necessary for the organism are also briefly described.展开更多
文摘The DNasel hypersensitive site 2 (HS2) of human β-globin locus control region (LCR) is required fOr the high level expression of human d-globin genes. In the present study, a stage-specific protein factor (LPF-β) was identified in the nuclear extract prepared from mouse fetal liver at d 18 of gestation, which could bind to the HS2 region of humanβ-globin LCRt We also found that the shift band of LPF-βfactor could be competed by humanβ-globin promoter. However, it couldn’t be competed by human E-globin promoter or by human Aβ-globin promoter. Furthermore, our data demonstrated that the binding-sequence of LPF-d factor is 5’CACACCCTA 3’,which is located at the HS2 region ofβ-LCR (from -10845 to -10853 bp) and humanβ-globin promoter (from -92 to -84 bp). We speculated that these regions containing the CACCC box in both the humallβ-globin promoter and HS2 might function as stage selector elements in the regulation of humanβd-globin switching and the LPF-βfactor might be a stage-specific protein factor involved in the regulation of humanβ-globin gene expression.
文摘In order to elucidate the molecular mechanisms of globin gene expression during embryonic development, the nuclear extracts from mouse hematopoietic tissue at different stages of development have been prepared. By using DNase I footprinting and gel mobility shift assays, the binding of protein factors in these extracts to the human βglobin promoter was analyzed. The differences in the binding patterns of protein factors during development were observed. An erythroid-specific and stage-specific nuclear protein in the nuclear extract from d 18 mouse fetal liver was identified, which can bind to the sequence (from -66bp to -90bp) of human β-globin promoter. We therefore speculate that the function of this cis-acting element may be similar to stage selector element (SSE) in chieken βA- promoter.
文摘Objective To investigate the effect of trans-acting factor(s) on rat glutathione S-transferase P1 gene (rGSTP1) transcription regulation in tumor cells.Methods The binding of trans-acting factor(s) to two enhancers of the rGSTP1 gene, glutathione S-transferase P enhancer Ⅰ (GPEI) and glutathione S-transferase P enhancer Ⅱ-1 (GPEⅡ-1), was identified by an electrophoretic mobility shift assay (EMSA). The molecular weight of trans-acting factor was measured in a UV cross-linking experiment. Results Trans-acting factor interacting with the core sequence of GPEI (cGPEI) were found in human cervical adenocarcinoma cell line (HeLa) and rat hepatoma cell line (CBRH7919). These proteins were not expressed in normal rat liver. Although specific binding proteins that bound to GPEⅡ-1 were detected in all three cell types, a 64 kDa binding protein that exists in HeLa and CBRH7919 cells was absent in normal rat liver. Conclusion cGPEI, GPEII specific binding proteins expressed in HeLa and CBRH7919 cells may play an important role in the high transcriptional level of the rGSTP1 gene in tumor cells.
基金supported by the National Natural Science Foundation of China(31270852,31000355)the National Key Basic Research Program of China(2012CB911000)
文摘Transfer RNA plays a fundamental role in the protein biosynthesis as an adaptor molecule by functioning as a biological link between the genetic nucleotide sequence in the mRNA and the amino acid sequence in the protein.To perform its role in protein biosynthesis,it has to be accurately recognized by aminoacyl-tRNA synthetases(aaRSs)to generate aminoacyl-tRNAs(aa-tRNAs).The correct pairing between an amino acid with its cognate tRNA is crucial for translational quality control.Production and utilization of mis-charged tRNAs are usually detrimental for all the species,resulting in cellular dysfunctions.Correct aa-tRNAs formation is collectively controlled by aaRSs with distinct mechanisms and/or other trans-factors.However,in very limited instances,mis-charged tRNAs are intermediate for specific pathways or essential components for the translational machinery.Here,from the point of accuracy in tRNA charging,we review our understanding about the mechanism ensuring correct aa-tRNA generation.In addition,some unique mis-charged tRNA species necessary for the organism are also briefly described.
