共查询到20条相似文献,搜索用时 62 毫秒
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Mouse, rat and human protein factors recognizing regulatory elements of nontranscribed spacer of rat ribosomal genes were studied by gel retardation assay. Protein factors bind specifically to the DNA fragments containing the core promoter sequence of RNA-polymerase I, to "spacer" promoter and to a putative enhancer sequence. Factors of mouse, rat and human nuclear extracts that recognize the region containing the core promoter sequence have similar molecular masses and are not identical to the previously described protein factor TIF-1B. Two factors that bind the "spacer" promoter region differ from the factors of the core promoter. "Spacer" promoter factors of mouse and rat nuclear extracts are probably identical, but differ from those of human extract. Protein factors, recognizing the putative enhancer region of rat and human extracts are alike but were not detected in mouse extract. Regions of nontranscribed spacer containing dispersed and tandem repeats do not bind any specific protein factors. 相似文献
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Isolation and characterization of a protein fraction that binds to enhancer core sequences in intracisternal A-particle long terminal repeats 总被引:5,自引:0,他引:5
The U3 region of mouse intracisternal A-particle (IAP) long terminal repeats (LTRs) contains several nuclear protein-binding domains. Two of these contain sequences with homology to the SV40 enhancer core. We refer to these two domains as Enh1 and Enh2. The Enh2 domain is an important determinant of promoter activity in vivo. We report here the isolation of nuclear fractions from human 293 and mouse MOPC-315 cells which interact with Enh1 and Enh2. Purification was achieved via DNA-affinity chromatography on a multimerized oligonucleotide representing the Enh2 region from the LTR of the mouse genomic IAP element, MIA14. Glycerol gradient sedimentation suggested a native Mr of approximately 80-100 for the binding component(s) in both crude and affinity-purified fractions. UV cross-linking showed that the binding activity involved two polypeptides within this size range. The affinity-isolated fraction from each cell line was highly purified, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and in vitro binding analysis. Exonuclease III footprinting showed that the two polypeptides interacted preferentially with the Enh1 and Enh2 domains within a 139-base pair segment from the MIA14 LTR. The polypeptides interacted in a sequence-specific manner with oligonucleotides representing these domains within the IAP LTR and with oligonucleotides containing the enhancer core sequence from SV40 and polyoma virus. Equilibrium binding studies indicated that the apparent dissociation constants for the polypeptides binding to the enhancer core sequence from MIA14, SV40, and polyoma virus were similar. Therefore, this affinity-purified fraction may represent a novel enhancer core-binding component which is distinct from the previously characterized rat CCAAT/enhancer-binding protein, C/EBP. 相似文献
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In order to further characterize the previously observed disruptive effect of the RNA polymerase I promoter sequence (Pol
I) from Acanthamoeba castellanii on tandemly repeated 5S rDNA positioning sequences from sea urchin (Lytechinus variegatus), we compared the histone-binding ability of the isolated 199-bp Pol I promoter region to that of the 208-bp 5S rDNA and
that of nucleosome core particle sequences isolated from chicken erythocytes. We found the 5S rDNA positioning sequence to
be more efficient at forming nucleosomes than the RNA polymerase I promoter sequence. Nevertheless, examination of the free-DNA
half-depletion points during the titrations suggested that twice as much histone had bound to RNA polymerase I promoter sequence
as to the 5S nucleosome-positioning or core particle sequences. DNA bending analysis suggested two potential DNA bending loci
in the RNA polymerase I promoter, whereas only one such locus was predicted for the 5S positioning sequence. Such mixed bending
signals on the RNA polymerase I promoter could favor non-nucleosomal deposition of histones on these sequences. 相似文献
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Expression of L-type calcium channels in cardiac myocytes and vascular smooth muscle cells (VSMC) critically regulates the contractile state of these cells. In order to discover the elements in the promoter region of the Ca(v)1.2 gene encoding the vascular/cardiac calcium channel alpha(1C) subunit that are important for the basal gene expression, approximately 2 kb of the 5'-flanking sequence of the Ca(v)1.2 gene has been cloned in our lab. In this study, using various lengths of the 5'-flanking DNA fused with a luciferase gene as a reporter, we have defined a 493-bp fragment of the cis-regulatory DNA which carries the majority of promoter activity in pulmonary artery smooth muscle (PAC1) cells. DNase I footprinting analysis of this 493-bp DNA using nuclear extracts from PAC1 cells revealed a 27-bp DNA sequence that contains a c-Ets like motif (CAGGATGC). Mutation of the Ets-like site and the respective flanking sequence within the DNase I footprinting protection region induced a marked change in the promoter activity in PAC1 cells. Electrophoretic mobility shift assays (EMSA) confirmed the presence of specific binding factor(s) in PAC1 cells' nuclear extracts for this 27-bp DNA. Competition studies with the wild-type and mutated DNA fragments established the importance of the 27 bp DNA sequence for high-affinity binding of the nuclear proteins to the promoter. We conclude that there is a 27 bp region in the promoter of the Ca(v)1.2 gene to which nuclear proteins from VSMC bind and strongly regulate the basal promoter activity. 相似文献
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Identification and characterization of a novel Schwann and outflow tract endocardial cushion lineage-restricted periostin enhancer 总被引:3,自引:1,他引:2
Lindsley A Snider P Zhou H Rogers R Wang J Olaopa M Kruzynska-Frejtag A Koushik SV Lilly B Burch JB Firulli AB Conway SJ 《Developmental biology》2007,307(2):340-355