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Query: UNIPROT:P06889 (Mol)
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G1 cyclins and some cyclin-dependent kinases (cdks) have been implicated in the G1/S transition during the eukaryotic cell cycle initiation. The present study demonstrates that the genes of cyclin E, cdk2, cdk5 and the transcriptional factor E2F-1 are expressed during the prolactin (PRL)-induced G1/S transition in rat Nb2 pre-T lymphoma cells. The mRNAs for these four cell cycle regulators were synergistically synthesized and degraded after the stimulation by PRL. The maximal levels of these mRNAs were observed at 8 to 12 h after the PRL addition, while DNA replication reached to the maximum between 12 and 16 h. These results suggest that cyclin E, cdk2, cdk5 and E2F-1 play the roles in the G1/S transition being expressed by a common cellular mechanism(s) in the PRL-stimulated pre-T lymphoma cells.
Biochem Mol Biol Int 1995 Oct
PMID:Synergistic gene expressions of cyclin E, cdk2, cdk5 and E2F-1 during the prolactin-induced G1/S transition in rat Nb2 pre-T lymphoma cells. 867 24

The early growth response-1 (Egr-1) gene has been identified as a nuclear transcriptional factor and implicated in the regulation of growth and differentiation of osteoblastic cells. In the present study, we investigated whether Egr-1 mRNA is expressed and induced by interleukin-1 beta, (IL-beta) and tumor necrosis factor-alpha (TNF-alpha) in normal human bone marrow stromal (HBMS) and osteoblastic (HOB) cells. Results demonstrate a very low basal expression of Egr-I mRNA which is induced by IL-1 beta, and TNF-alpha in a time- and dose-dependent manner. Egr-1 mRNA induction was detectable within 15 min, reached maximal by 60 min and thereafter declined to basal levels by 120 min. Induction of Egr-1 mRNA by IL-1 beta and TNF-alpha was completely inhibited by H-7 suggesting the mediation of protein kinase C. The induction by IL-1 beta and TNF-alpha of Egr-1 mRNA was independent of de novo protein synthesis since this induction was also observed in the presence of protein synthesis inhibitor cycloheximide. Fetal bovine serum and cycloheximide also independently induced the Egr-1 mRNA. Actinomycin D experiments demonstrated that Egr-1 mRNA is degraded very rapidly with a half-life of 30 min. Our results demonstrate the expression of Egr-1 gene and its induction by IL-1 beta, and TNF-alpha in normal human bone marrow stromal (osteoprogenitor) and osteoblastic cells in primary cultures. Data also reveal that the expression of Egr-1 gene is inhibited by protein kinase C inhibitor H-7 suggesting that the activation of protein kinase C or other protein kinases resulting in the phosphorylation of specific transcription factor(s) is the first immediate early step in the induction of immediate-early Egr-1 gene by IL-1 beta, and TNF-alpha. Results also suggest that Egr-1 is an important mediator of IL-1 beta and TNF-alpha action in normal human osteoblastic cells.
Mol Cell Biochem 1996 Mar 09
PMID:Induction of early growth response-1 gene by interleukin-1 beta and tumor necrosis factor-alpha in normal human bone marrow stromal an osteoblastic cells: regulation by a protein kinase C inhibitor. 870 78

Using electrophoretic mobility-shift assay (EMSA), we examined changes in DNA-binding activities of transcriptional factor-activated protein-1 (AP-1), which is a Fos-Jun protein complex, onto its responsive element TRE in the hippocampus and amygdaloid nucleus of rats stimulated with pentylenetetrazol (PTZ) injection, and also investigated the effects of a single administration of the immunosuppressant cyclosporin A (CsA). In EMSA with nuclear extracts from the rat brain, the TRE-binding activity of AP-1 in the hippocampus and amygdaloid nucleus markedly increased 2 h after the PTZ injection (75 mg/kg, i.p.). These PTZ-induced increases of the TRE-binding protein in these regions were completely suppressed, by pretreatment with CsA (5 mg/kg, s.c.) 1 h before the PTZ injection. In addition, the administration of CsA significantly ameliorated PTZ-induced convulsion. This therapeutic effect of single CsA pretreatment may be based, in part, on the effects on the TRE-binding activity of AP-1 in the brain. Since single pretreatment of CsA in the present study had no effect on the PTZ-induced induction of c-fos mRNA, c-jun mRNA, Fos protein nor Jun protein, the inhibitory effects of single CsA administration on PTZ-induced TRE-binding activity in the brain may be related to the effects of CsA on AP-1 itself. These results suggest that an immune response via activation of transcriptional factor in the brain tissue is involved in the convulsion.
Brain Res Mol Brain Res 1995 Oct
PMID:Effects of single cyclosporin A pretreatment on pentylenetetrazol-induced convulsion and on TRE-binding activity in the rat brain. 877 43

We cloned novel cDNAs from MB02 human erythroleukemia cells using PCR based approaches: a) Differential display by means of RT-PCR using one 5' primer CTTGATTGCC and four different 3' primers (T12AA, T12CA, T12GA, and T12AT). Ninety-three percent of the differential clones which were reamplified and sequenced were cDNAs of previously unidentified genes. b) Cloning using degenerate TFIIIA-like zinc finger domain specific oligonucleotide. Of the 54 clones sequenced, 20 contained two or more zinc finger sequences. Ten of these clones were new zinc finger cDNAs and one belonged to a known zinc finger gene (ZFP7). c) Cloning using degenerate tyrosine kinase(TK) domain-specific oligonucleotides corresponding to the highly conserved amino acid sequences IHRDLAA and DVWSFG. Of the 28 cDNA clones sequenced, 7 were JAK1 TK, one was atk TK, one was tec TK. The remaining sequences belonged to new ESTs or to ribosomal genes. d) Cloning using degenerate POU domain-specific oligonucleotides corresponding to sequence FK(QV)RRIK of the POU-specific domain and to sequence VWFCN(RQ)R of the POU-homeodomain. Sixteen clones were sequenced and all but one were identical with the Oct-1 transcriptional factor. Differential display RT-PCR and PCR-based cDNA cloning using degenerate primers for zinc finger motifs yielded the largest number of new genes expressed in MBO2 cells.
Blood Cells Mol Dis 1996
PMID:Identification of new genes expressed in a human erythroleukemia cell line. 880 82

The product of the Escherichia coli aidB gene is homologous to human isovaleryl-coenzyme A dehydrogenase (IVD), an enzyme involved in the breakdown of the amino acid leucine. The aidB gene is not expressed constitutively, but its transcription is induced via distinct mechanisms in response to: (i) exposure to alkylating agents; (ii) acetate at a slightly acidic pH; and (iii) anoxia. Induction by alkylating agents is mediated by the transcriptional activator Ada, in its methylated form (meAda); the other forms of induction are Ada independent and require sigma s, the alternative sigma factor mainly expressed during the stationary phase of bacterial growth. In this report we show that, in the absence of any transcriptional factor, aidB is efficiently transcribed in vitro by the sigma s, but not by the sigma 70, form of RNA polymerase holoenzyme. In the presence of meAda, levels of transcription by both forms of RNA polymerase are significantly increased. However, sigma s-dependent transcription of aidB is inhibited both in vitro and in vivo by binding of the transcriptional regulator Lrp (leucine responsive protein) to the aidB promoter region (PaidB). Lrp acts as a specific repressor for sigma s-dependent transcription of aidB. Leucine counteracts Lrp binding to P aidB, as does binding to P aidB of me Ada, which causes Lrp to dissociate from the promoter. The physiological significance of aidB transcription regulation is discussed.
Mol Microbiol 1996 Jun
PMID:The leucine-responsive regulatory protein (Lrp) acts as a specific repressor for sigma s-dependent transcription of the Escherichia coli aidB gene. 880 48

Primers were developed to allow the rapid and reliable assay of heat shock transcriptional factors 1 and 2 in human epidermoid A431 cells by following the protocol described in this study. Using the primers, the heat-induced increase in heat shock transcriptional factor 1 but not 2 was observed. This is the first report to show that heat shock increases the mRNA amount of HSF1 with no changes in HSF2 mRNA.
Mol Cell Biochem 1996 May 24
PMID:Rapid assay of HSF1 and HSF2 gene expression by RT-PCR. 881 81

Expression of the outer membrane protein OmpF of Escherichia coli K-12 is influenced by a variety of environmental signals. Most of the signals are thought to regulate OmpF expression at the level of transcription initiation. A key element of this regulation is the interaction between the transcriptional factor OmpR and the cis-acting regulatory region of ompF. In this study, we used a combination of DNase I, dimethyl sulfate and hydroxyl radical footprinting analysis and DNA migration retardation assays to identify the bases within the ompF regulatory region that are in contact with OmpR. Our results indicate that the -107 to -39 region of ompF contains three individual binding sites and that a single OmpR-binding site is capable of interacting with two OmpR molecules. We also establish that a single OmpR-binding site is composed of two half-sites and that both half-sites are required for the formation of stable OmpR/DNA complexes. Comparisons of the sequences protected by OmpR indicate that an OmpR-binding site spans approximately 18 bp and has two highly conserved G/C base-pairs that are separated by three nucleotides. Although the three OmpR-binding sites we identified exhibit limited sequence similarity, this may reflect the fact that two of the sites are incapable of binding OmpR independently and can bind OmpR only if adjacent to another OmpR-binding site. Finally, our DNA migration retardation assays suggest that phosphorylation stimulates the cooperative interactions between OmpR molecules bound at neighboring sites. Therefore, this study provides a detailed understanding of how OmpR interacts with its binding sites immediately upstream of ompF and serves as a foundation for studying how phosphorylation of OmpR results in the regulation of ompF expression in response to environmental signals.
J Mol Biol 1996 Oct 11
PMID:Identification of the bases in the ompF regulatory region, which interact with the transcription factor OmpR. 887 42

To gain insight into the interactions between transcriptional factor proteins and DNA, the DNA-reactive drugs (+)-CC-1065 and pluramycin were used to target specific protein-DNA complexes. The structural features of the complex between the transcriptional activator Sp1 and the 21-base-pair repeat of the early promoter region of SV40 DNA were examined using hydroxyl-radical footprinting; (+)-CC-1065, a sequence-specific minor groove bending probe; and circularization experiments. The results show that the 21-base-pair repeat region has an intrinsically in-phase bent structure that is stabilized upon saturation Sp1 binding by protein-DNA and protein-protein interactions to produce a looping structure. The intercalating drug pluramycin was used to probe the structural details of the interaction between the TATA binding protein (TBP) and the TATA box DNA sequence. TBP, which directs initiation of RNA transcription, exhibits two-fold symmetry and apparently interacts with the TATA box in a symmetrical fashion. However, the interaction results in an asymmetric effect, in that transcription is initiated only in the downstream direction. Using pluramycin as a probe, it was determined that TBP binding to the human myoglobin TATA sequences enhances pluramycin reactivity at a site immediately downstream of the TATA box. The implications on transcriptional control of ternary complexes comprised of transcriptional factors, DNA, and DNA-reactive compounds will be presented.
J Mol Recognit
PMID:Specific targeting of protein-DNA complexes by DNA-reactive drugs (+)-CC-1065 and pluramycins. 887 97

The homeodomain protein PDX-1, referred as IPF-1/STF-1/IDX-1, is a transcriptional factor that plays a critical role in the control of several genes expressed in the pancreatic islet. PDX-1 gene expression has been previously shown to be reduced in cultured beta-cell lines chronically exposed to high glucose concentrations. As the glucose transporter type 2 (GLUT2) gene expression is selectively decreased in the beta-pancreatic cells of experimental models of diabetes, we postulated that the loss of GLUT2 gene expression in the pancreatic islets of diabetic animals may be due to the loss of PDX-1 transacting function on the GLUT2 gene. We, therefore, investigated the potential role of PDX-1 in the transcriptional control of GLUT2. We have identified a repeat of a TAAT motif (5'-TAATA-ATAACA-3') conserved in the sequence of the human and murine GLUT2 promoters. Recombinant PDX-1 binds to this GLUT2TAAT motif in electrophoretic mobility shift experiments. PDX-1 antiserum detects the formation of the complex of PDX-1 with the GLUT2TAAT motif in nuclear extracts from the pancreatic insulin-secreting cell line, beta TC3. The GLUT2TAAT motif was mutated in the murine GLUT2 promoter (-1308/+49 bp) linked to a luciferase reporter gene and transfected into beta TC3 cells. Compared with the transcriptional activity of the wild type promoter, that of the mutated promoter decreases by 41%. Multiple copies of the GLUT2TAAT motif were ligated 5' to a heterologous promoter and transfected into a PDX-1-expressing cell line (beta TC3) and into cell lines lacking the homeobox factor (InR1-G9 and JEG-3). The GLUT2TAAT motif mediates the activation of the heterologous promoter in the PDX-1-expressing cell line but not in InR1-G9 or JEG-3 cell lines. Furthermore, cotransfection in a PDX-1-deficient cell line with the expression vector encoding PDX-1 transactivates specifically the heterologous promoter containing the multimerized GLUT2TAAT motif. These data demonstrate that the murine GLUT2 promoter is controlled by the PDX-1 homeobox factor through the identified GLUT2TAAT motif.
Mol Endocrinol 1996 Nov
PMID:Transcriptional activation of the GLUT2 gene by the IPF-1/STF-1/IDX-1 homeobox factor. 892 59

In an initial effort to understand the molecular mechanism of how low temperature induces sorbitol dehydrogenase gene expression in diapause eggs of the silkworm, the sorbitol dehydrogenase gene was isolated from a Bombyx genomic library using a cDNA encoding the Bombyx homologue of mammalian sorbitol dehydrogenase as a probe. The gene extended for about 10 kb, consisting of eight exons and seven introns. Four TATA motifs were found in the 5' upstream region of the gene, without CCAAT. AATTAA, instead of AATAAA, was localized in the upstream region of the polyadenylation site. Although a single copy of this gene was present per haploid genome, 1.2 kb and 1.1 kb transcripts were found from yolk cells in diapause eggs and from larval fat-body cells, respectively. The two major transcription initiation sites corresponding to both transcripts were localized at 355 and 226 base pairs upstream from the transition start site, indicating an alternative use of promoter. The 5'-upstream region of the gene contained a consensus sequence, TGA(A/T)AA(A/G/T), that has been found in insect genes expressed mainly in larval and pupal fat bodies. It also contained three kinds of sequences similar to cis-elements recognized by members of the steroid receptor superfamily, such as chicken ovalbumin upstream promoter transcription factor (COUP-TF)/Drosophila Seven up (SVP), Drosophila hormone receptor 39 (DHR39) and Bombyx fushi tarazu transcriptional factor 1 (BmFTZ-F1).
Insect Mol Biol 1996 Nov
PMID:Structure of the Bombyx sorbitol dehydrogenase gene: a possible alternative use of the promoter. 893 78


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