Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.3.1.28 (chloramphenicol acetyltransferase)
5,100 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human c-myb is normally involved in the regulation of proliferation and differentiation of hematopoietic cells. Until now, only a few reports have described elevated c-myb gene expression in epithelial tissue, suggesting that under certain circumstances, c-Myb protein might play a role during the process of malignant transformation of epithelial cells. To investigate a possible role of c-myb during papillomavirus-associated carcinogenesis, we investigated the c-myb mRNA expression in human papillomavirus (HPV)-associated tumors and tumor cell lines. Seven of nine cervical carcinomas and two of three carcinoma cell lines exhibited elevated c-myb transcriptional activity. In contrast to malignant cervical neoplasias, only 3 of 15 condylomata acuminata expressed a sparse signal for c-myb mRNA. Since the c-Myb protein has been described as a potent transcriptional regulator, we investigated the transactivating properties of c-Myb on the HPV-16 promoter/enhancer. Cotransfection of a chloramphenicol acetyltransferase-reporter plasmid containing the HPV-16 enhancer/promoter element with a full-length c-Myb-expressing plasmid resulted in a significant induction (4.3-fold) of the HPV-16 promoter, whereas expression of a carboxy-terminally deleted c-Myb protein led to no effects. Gel shift experiments showed a specific binding of recombinant c-Myb protein on the HPV-16 P97 enhancer. These data indicate that elevated c-myb expression occurs with HPV-associated cell transformation. Since c-Myb has been shown to stimulate the HPV-derived oncoprotein expression via transcriptional activation, it may play a role in the process of HPV-associated cervical carcinogenesis.
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PMID:Human c-myb is expressed in cervical carcinomas and transactivates the HPV-16 promoter. 767 Dec 56

Since we have observed effects of growth factors and cAMP as well as estradiol (E2) on regulation of expression of some genes stimulated by the estrogen receptor (ER), we have undertaken studies to examine directly whether activators of protein kinases can modulate transcriptional activity of the ER. We find that activators of protein kinase-A [cholera toxin plus 3-isobutyl-1-methylxanthine (CT+IBMX)] and protein kinase-C [12-O-tetradecanoylphorbol-13-acetate (TPA)] markedly synergize with E2 in ER-mediated transcriptional activation. When a reporter plasmid [with a minimal promoter containing a TATA region and estrogen-responsive elements (ERE) linked to a chloramphenicol acetyltransferase (CAT) reporter gene] was transfected into MCF-7 human breast cancer cells, which contain endogenous ER, E2 evoked a dose-dependent increase in CAT activity. While treatment with protein kinase-A or -C activator alone evoked only very low CAT activity, the maximal (approximately 25-fold) CAT activity stimulated by E2 alone was increased 2- to 3-fold (to approximately 60 times the control value) upon cotreatment with either of the protein kinase activators. Interestingly, antiestrogen abolished all of the CAT activity induced by E2 and protein kinase activators. Immunoblots showed that TPA reduced ER levels to 30% of control values after 24 h, while CT+IBMX increased levels about 1.5-fold. Scatchard binding analysis revealed no change in the binding affinity of E2 to ER by these agents. Gel mobility shift competition assays with extracts prepared from cells that had been treated with E2 and protein kinase activators did not reveal any quantitative or qualitative changes in the binding of ER to the ERE in vitro. In ER-deficient Chinese hamster ovary (CHO) cells transfected with the reporter gene and varying amounts of an ER expression vector, the level of CAT activity obtained by cotreatment with E2 and CT+IMBX was 3-fold higher than that observed with E2 alone over the range of different ER amounts tested. This ER-mediated synergism was still retained in an amino-terminal A/B-domain-deleted ER mutant lacking the hormone-independent transcriptional activation function (TAF-1), but was greatly reduced in two hormone-binding domain (region E) mutants that exhibit significantly diminished ligand-dependent transcriptional activation. TPA did not show any synergistic activation with E2 in CHO cells, indicating differences in responses between cell types.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Synergistic activation of estrogen receptor-mediated transcription by estradiol and protein kinase activators. 768 75

Insulin-like growth factor binding protein-1 (IGFBP-1) is expressed primarily in the liver, kidney, and uterus. Basal IGFBP-1 promoter activity in human HEP G2 hepatoma cells is dependent upon a proximal promoter element that binds hepatic nuclear factor 1 (HNF1), a protein that is likely to be an important factor regulating the expression of many genes in liver and kidney. To test whether HNF1 activates IGFBP-1 transcription, HEP G2 cells and HeLa cells were cotransfected transiently with HNF1 expression vectors and with IGFBP-1 promoter/chloramphenicol acetyltransferase reporter gene constructs. HNF1 increased IGFBP-1 promoter activity in both HEP G2 and HeLa cells. Gel mobility-shift assays and additional transfections in HeLa cells showed that expressed full-length and carboxy-terminal truncated forms of HNF1 could each bind the HNF1 cis element of the IGFBP-1 promoter; however, significant trans-activation only occurred in the presence of the full-length HNF1 protein, similar to past experience with these two HNF1 forms and the albumin promoter. Further studies showed that IGFBP-1 promoter constructs containing mutations with high or low affinity for HNF1 responded to HNF1 expression with increased or decreased activity, respectively, relative to the native promoter. These studies suggest that HNF1 and/or related proteins play a role in hepatic, and perhaps also renal, expression of IGFBP-1.
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PMID:HNF1 activates transcription of the human gene for insulin-like growth factor binding protein-1. 768 29

We have examined mechanisms of regulation of the human glycoprotein hormone alpha subunit gene by thyroid hormone (T3) and estradiol. Pituitary-derived GH3 cells were transiently transfected with chimeric constructs comprising between 1,500 and 98 base pairs of human alpha subunit gene 5'-flanking sequence fused to the bacterial gene encoding chloramphenicol acetyltransferase (h alpha CAT) and treated with T3 and estradiol, alone and in combination. In pituitary cells, 98 base pairs of alpha gene 5'-flanking sequence were sufficient to mediate both inhibition of alpha gene promoter activity by T3 and stimulation by estradiol; inhibition of the alpha promoter by T3 was antagonized by estradiol. Mutation of nucleotides essential for T3 receptor binding to the alpha gene thyroid hormone response element abolished the response of h alpha CAT expression to estradiol as well as T3. In contrast to pituitary GH3 cells, estradiol treatment alone had no effect on expression of either h alpha CAT or the endogenous alpha gene in JEG-3 choriocarcinoma cells cotransfected with a human thyroid hormone receptor expression vector, but estradiol antagonized suppression of both endogenous and transfected alpha promoter activity by T3. Gel mobility shift assays demonstrated specific binding of in vitro synthesized human estrogen receptor (ER) to the alpha gene thyroid hormone response element. These findings suggest that estradiol modulates expression of the human alpha subunit gene in pituitary and choriocarcinoma cells by direct binding of ER to the alpha gene promoter, and that interaction of ER with the alpha gene negative TRE accounts for the antagonistic effects of estradiol and T3.
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PMID:Estradiol modulates thyroid hormone regulation of the human glycoprotein hormone alpha subunit gene. 769 20

The androgen receptor (AR) is a nuclear transcription factor that is essential for development of the male urogenital tract. In the current work, we have characterized the mouse androgen receptor suppressor (mARS). A single, 20-base pair, region (TCCCCCCACCCACCCCC-CCT) was sufficient for suppression in chloramphenicol acetyltransferase assays. Northern analysis indicated that translational regulation is not necessary for the suppression. Analysis of the AR mRNA half-life indicated that the mARS does not affect AR RNA degradation. Gel mobility assays showed that the mARS is bound by multiple proteins that can recognize single-stranded DNA and RNA. In addition, differing proteins are expressed in distinct tissues. Purification of some of these proteins has shown that a doublet of 33 and 35 kDa binds to the G-rich strand and that a 52-kDa protein binds to the C-rich strand. Southwestern blots have confirmed that these proteins are indeed recognized by the mARS. The results of these experiments indicate that the AR 5'-untranslated region contains a suppressor element that can be bound by multiple proteins. The mARS appears to be acting either by altering transcription initiation or blocking transcription elongation. Characterization of this suppressor may provide insight into the physiological means by which the AR is regulated.
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PMID:The androgen receptor is transcriptionally suppressed by proteins that bind single-stranded DNA. 773 38

Mouse calbindin-D28k expression is regulated in vivo by estradiol in ovaries, uterus, and oviduct. To determine whether estrogen can have an effect on the transcription of the calbindin-D28k gene, the human breast cancer cells T47D were transiently transfected with a plasmid containing a 1.1 kilobase (kb) PstI/SacII fragment (-1075/+34) of the mouse calbindin gene ligated to the chloramphenicol acetyltransferase (CAT) gene and cotransfected with human estrogen receptor expression vector. T47D cells, transfected and treated with estradiol (10(-11) - 10(-7) M for 64-65 h), exhibited a dose-dependent increase in CAT activity (up to 6.2-fold). Transfection of MCF-7 breast cancer cells with the chimeric gene construct alone also resulted in an estradiol-dependent induction in CAT activity. Deletion mutant analysis demonstrated that there are two regions of the mouse calbindin-D28k promoter (between -1075/-702 and between -175/-78) that contribute to the induction by estradiol. These fragments, when linked to the thymidine kinase promoter to construct a heterologous promoter chimera, were able to convert the thymidine kinase promoter to estrogen responsiveness. In these regions there are multiple imperfect half-palindromic estrogen-responsive elements. Gel retardation assays demonstrated weak protein-DNA interactions that were competed with cold oligonucleotide containing the vitellogenin estrogen-response element. These findings indicate that the mouse calbindin-D28k promoter is capable of conferring estrogen responsiveness, which may be mediated by several imperfect half-palindromic estrogen-responsive elements, and suggest, in light of previous studies concerning 1,25-dihydroxyvitamin D regulation, multiple steroid regulation of the calbindin-D28k gene.
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PMID:Regulation by estrogen through the 5'-flanking region of the mouse calbindin-D28k gene. 777 78

A mesenchymal-epithelial cell interaction exists in the testis between the Sertoli cells that form the seminiferous tubule and the mesenchymal-derived peritubular myoid cells that surround the tubule. Analysis of the mesenchymal-epithelial interactions between these cells revealed the local production of a mesenchymal factor, PModS. PModS modulates the differentiated functions of Sertoli cells in vitro, including stimulation of the iron-binding protein transferrin (Tf). Previous results have indicated that PModS-induced Tf gene expression involves the activation of immediate early genes. One of the immediate early genes was identified as c-fos. The importance of c-fos was demonstrated in the current study when a c-fos antisense oligonucleotide was found to inhibit the ability of PModS to induce the expression of a Tf promoter-chloramphenicol acetyltransferase (CAT) construct. The regulation of c-fos by PModS was investigated with various CAT constructs containing segments of the c-fos promoter, such as the serum response element (SRE), sis-inducible element (SIE), cAMP response element (CRE), and phorbol ester/TPA response element (TRE), transfected into cultured Sertoli cells. PModS has no effect on cAMP response element-CAT or TRE-CAT, suggesting that PModS does not act through stimulation of cAMP and protein kinase C pathways. PModS was found to activate the c-fos SRE-CAT construct and the SIE-CAT construct. A construct containing both SIE and SRE was stimulated to the same degree as either element alone. Gel mobility shift assays using nuclear extracts from PModS-stimulated Sertoli cells and a radiolabeled SRE oligonucleotide resulted in retarded mobility of a DNA-protein complex. A gel shift with a SRE oligonucleotide containing an ETS domain resulted in a unique shift only detected in PModS stimulated cells. PModS also promoted a gel shift with the SIE that is adjacent to the SRE on the c-fos promoter. The data imply that PModS can activate the c-fos promoter through the SRE and SIE. PModS caused a labeled activating protein 1 (AP1) oligonucleotide to form a DNA-protein complex, indicating activation of the c-fos gene and binding of the c-fos/jun complex. To study the downstream regulation of Sertoli cell differentiation, Tf gene expression was examined. CAT constructs containing deletion mutants of a 3-kilobase (kb) mouse Tf promoter were used. When transfected into Sertoli cells the 581-base pair Tf minimal promoter had only a slight response to PModS, but was activated by FSH. The 2.6-kb Tf promoter construct responded to PModS.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Role of specific response elements of the c-fos promoter and involvement of intermediate transcription factor(s) in the induction of Sertoli cell differentiation (transferrin promoter activation) by the testicular paracrine factor PModS. 778 31

Transfection experiments with constructs containing various 5'-deleted fragments of the human lipoprotein lipase (LPL) promoter and the chloramphenicol acetyltransferase reporter gene revealed an LPL silencer element (LSE) in the region of nucleotides -225 to -81 of the LPL gene that functioned in Chinese hamster ovary (CHO) and HeLa cells. Gel retardation competition analysis showed the presence of a nuclear factor(s) capable of binding to the sequence of nucleotides -169 to -152 of LSE (LSE-6) in a single-stranded (opposite-strand) and double-stranded specific fashion, the binding affinity being almost the same in the two binding forms. Site-directed mutagenesis indicated that almost the entire sequence of LSE-6 was necessary to form the complexes and also critical for silencing activity in CHO cells. The amounts of this binding factor(s) in CHO and HeLa cells were closely associated with transcriptional silencing activity. Photochemical cross-linking experiments indicated that the single- and double-stranded elements recognized the same binding factor(s) with molecular masses of 54 to 63 kDa and 109 to 124 kDa. The 109- to 124-kDa DNA binding factor(s) was found to be a doublet of that of the 54- to 63-kDa factor by isoelectric focusing or by increasing the time of exposure to UV irradiation. When inserted upstream of another gene such as that of the simian virus 40 enhancer/promoter of pSV2CAT, the sequence of nucleotides -190 to -143 (LSE-1) also suppressed transcription of the reporter gene in CHO cells. These results strongly suggest that the LSE plays a role in regulation of LPL gene expression by suppressing its transcription.
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PMID:A silencer element for the lipoprotein lipase gene promoter and cognate double- and single-stranded DNA-binding proteins. 779 60

Previously, we reported that mesangial cells increased fibronectin, laminin and type IV collagen synthesis when cultured in the presence of high glucose (30 mM). Although mRNA levels for all three extracellular matrix (ECM) proteins were also increased in high glucose conditions, the mechanism for this increase was not known. In order to determine whether increased transcription was involved in the observed increase in fibronectin mRNA levels mesangial cells were transfected with a construct containing the 5'-flanking region of the fibronectin (FN) gene [position +69 to -510 base pairs (bp)] fused to the coding region of the chloramphenicol acetyltransferase (CAT) gene [FN-CAT (-510)]. Cells were transiently and stably transfected with this construct. Under serum-free conditions, high glucose increased CAT activity only in the presence of TGF beta 1 (referred to as TGF beta). The experiments were performed without serum because FN-CAT (-510) contains a serum responsive element. The increase in CAT was approximately twofold in transiently transfected cells and threefold in stably transfected cells. TGF beta alone increased CAT activity approximately 30%. Stimulation of fibronectin gene expression appeared to occur at the level of a cAMP response element (CRE) located -170 bp of the FN gene because cells transfected with a construct containing an oligonucleotide encoding for this CRE fused to a minimal fibronectin promoter (-56 bp) and a CAT reporter gene [CRE (-170) FN-CAT] displayed similar increments of CAT activity after treatment with high glucose and TGF beta. Gel shift mobility assays with a CRE oligonucleotide revealed multiple complexes with mesangial cell nuclear proteins.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:High glucose and TGF beta 1 stimulate fibronectin gene expression through a cAMP response element. 786 96

The functional induction of c-fos in the sodium butyrate-induced differentiation of F-98 glioma cells was studied. Fos protein level was increased by butyrate. In contrast, c-Jun protein was constitutively expressed and was not affected by butyrate. Gel-retardation assay indicates Fos as a component of the complex formed between the consensus oligonucleotide of the TPA (PMA, phorbol 12-myristate 13-acetate) response element (TRE) and nuclear extract prepared from butyrate-treated cells. Transfection studies showed that butyrate increased transcription from a multimeric TRE-driven reporter construct, and the effect was mimicked by transfecting cells with fos-expression plasmid. Furthermore, under conditions of c-fos over-expression, transactivation by butyrate was essentially abolished. These data suggest that Fos induction had a functional role in gene activation. Characterization of stable c-fos transfectants demonstrated that these cells displayed alterations in morphology, showed serum-dependent growth, had slower growth rates and grew to lower saturation densities than did untransfected F-98 cells or transfected cells that did not express c-fos. Immunofluorescent staining indicated that fos transfectants also had elevated glial fibrillary acidic protein ('GFAP') expression. Transfection of the c-fos promoter-chloramphenicol acetyltransferase fusion gene into F-98 cells revealed that activation of c-fos by butyrate was exerted at the promoter level, and sequences located within nucleotides -757 to -402 of the c-fos promoter were responsible for butyrate induction. Our data indicate that transcriptional activation of c-fos through its promoter by butyrate resulted in increased Fos protein expression. Transfection studies show that both c-fos and butyrate activate TRE-containing genes, and fos may be a downstream mediator of butyrate. Furthermore, expression of c-fos plays a major role in modulating the growth properties of F-98 cells.
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PMID:Analysis of c-fos expression in the butyrate-induced F-98 glioma cell differentiation. 786 28


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