Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

H4IIE rat hepatoma cells were stably transfected with various phosphoenolpyruvate carboxykinase-chloramphenicol acetyltransferase (PEPCK-CAT) expression vectors. The regulation of the transfected genes was qualitatively similar to that of the endogenous PEPCK gene. CAT expression was increased in response to cAMP and dexamethasone and insulin overrode these effects at concentrations known to be effective in suppressing transcription of the endogenous gene. The effect of insulin was dominant, as it is with the endogenous gene. A series of 5',3', and internal deletions of the PEPCK gene promoter were used to show that this insulin response requires at least two separate elements. One insulin-responsive sequence is located between -468 and -402, relative to the transcription initiation site. The other is between -271 and +69.
Mol Endocrinol 1990 Sep
PMID:Regulation of phosphoenolpyruvate carboxykinase gene expression by insulin. Use of the stable transfection approach to locate an insulin responsive sequence. 217 98

Full-length rat and human androgen receptor (AR) cDNA clones were expressed in COS-7 and CV1 monkey kidney cells to analyze the AR protein using immunological and cotransfection techniques. The studies were aided by the development of two rabbit polyclonal antibodies, designated AR32 and AR52, directed against epitopes within the N-terminal region of AR. Each antibody recognizes native AR by sucrose gradient analysis and detects a 114-kilodalton protein in COS cells transfected with human or rat AR cDNA. Covalent binding of the synthetic androgen [3H]methyltrienolone (R1881) to the 114-kDa protein was saturable. The endogenous native AR was similarly 114 kDa on immunoblots of a human prostate adenocarcinoma cell line, LNCaP, and rat sex accessory gland extracts. AR was localized in nuclei of transfected COS cells and in LNCaP cells by immunocytochemical staining. Androgen induction of CAT activity was dose dependent in CV1 cells cotransfected with the AR expression vector and a reporter plasmid containing the mouse mammary tumor virus promoter linked to the chloramphenicol acetyltransferase gene. It is concluded that antipeptide antibodies are useful reagents in characterizing both native and denatured forms of the AR protein. The 114-kDa protein expressed transiently in cultured cells represents the full-length AR protein, has a molecular size equivalent to that of endogenous AR, and mediates androgen-dependent transcriptional activation in CV1 cells.
Mol Endocrinol 1990 Sep
PMID:Expression of recombinant androgen receptor in cultured mammalian cells. 217 2

High level bacterial resistance to chloramphenicol is generally due to O-acetylation of the antibiotic in a reaction catalysed by chloramphenicol acetyltransferase (CAT, EC 2.3.1.28) in which acetyl-coenzyme A is the acyl donor. The crystal structure of the type III enzyme from Escherichia coli with chloramphenicol bound has been determined and refined at 1.75 A resolution, using a restrained parameter reciprocal space least squares procedure. The refined model, which includes chloramphenicol, 204 solvent molecules and two cobalt ions has a crystallographic R-factor of 18.3% for 27,300 reflections between 6 and 1.75 A resolution. The root-mean-square deviation in bond lengths from ideal values is 0.02 A. The cobalt ions play a crucial role in stabilizing the packing of the molecule in the crystal lattice. CAT is a trimer of identical subunits (monomer Mr 25,000) and the trimeric structure is stabilized by a number of hydrogen bonds, some of which result in the extension of a beta-sheet across the subunit interface. Chloramphenicol binds in a deep pocket located at the boundary between adjacent subunits of the trimer, such that the majority of residues forming the binding pocket belong to one subunit while the catalytically essential histidine belongs to the adjacent subunit. His195 is appropriately positioned to act as a general base catalyst in the reaction, and the required tautomeric stabilization is provided by an unusual interaction with a main-chain carbonyl oxygen.
J Mol Biol 1990 May 05
PMID:Refined crystal structure of type III chloramphenicol acetyltransferase at 1.75 A resolution. 218 98

The sequences preceding the albumin mRNA start site are able to direct efficient transcription only upon introduction into cells expressing the endogenous albumin gene. In transient expression assays, the activity of a reporter gene (CAT) linked to this promoter is 100-fold higher in H4II differentiated hepatoma cells than in H5 dedifferentiated cells which no longer express their albumin gene. This tissue specificity depends on the very proximal promoter region, composed of a CCAAT box, the proximal element and a TATA box. Deletion of the CCAAT box leads to a two- to threefold decrease in activity, deletion of the proximal element (PE) results in loss of activity. The PE is a high-affinity binding site for HNF1/APF, a strictly liver specific trans-acting factor. When the affinity of this factor for PE is decreased by bacterial methylation (PE includes a dam methylase site), by mutation, or by its replacement with the homologous element from the alpha-fetoprotein gene (AFP), the activity of the short promoter (PE plus the TATA box) is abolished. This activity can be rescued in the presence of the more upstream elements: DEII, DEI and the CCAAT box (recognized, respectively, by the NF1/CTF, C/EBP and NFY/ACF factors) which are then absolutely required. Our results suggest that the upstream elements contribute to promoter activity by stabilizing the HNF1-PE complex and not by direct interaction with TFIID or the RNA polymerase. It is probable that these elements, essentially dispensable in already differentiated hepatoma cells, play a crucial role during development or differentiation to activate the promoter in cells that contain a low concentration of HNF1 and/or an HNF1 unable to open inactive chromatin alone.
Mol Biol Med 1990 Apr
PMID:Anatomy of the rat albumin promoter. 218 62

Three DNA constructs, the natural human growth hormone gene (hGH-hGH) its 500 bp promoter linked to the chloramphenicol acetyl transferase reporter gene (hGH-CAT), and its structural part linked to the herpes virus thymidine kinase promoter (TK-hGH) were introduced into rat pituitary GC cells by DEAE-dextran transfection. Transient expression was followed as a function of triiodothyronine (T3) concentration. The hGH-CAT expression was specifically inhibited by T3 following a typical dose-response curve while hGH-GH gene expression was not significantly modified. The transient expression of TK-hGH increased as a function of T3 concentration. These results indicate that T3 exerts two opposite effects on hGH gene expression. First, it down-regulates expression by acting on the promoter; second, it up-regulates expression by acting on the structural part of the gene. These action could be due to regulation of transcription and mRNA stabilization, respectively.
Mol Cell Endocrinol 1990 Jul 09
PMID:Triiodothyronine inhibits transcription from the human growth hormone promoter. 221 33

The androgen receptor (AR) is activated upon binding of testosterone or dihydrotestosterone and exerts regulatory effects on gene expression in androgen target cells. To study transcriptional regulation of the rat AR gene itself, the 5' genomic region of this gene was cloned from a genomic library and the promoter was identified. S1-nuclease protection analysis showed two major transcription start sites, located between 1010 and 1023 bp upstream from the translation initiation codon. The area surrounding these start sites was cloned in both orientations in a CAT reporter plasmid. Upon transfection of the constructs into COS cells, part of the promoter stimulated transcription in an orientation-independent manner, but the full promoter showed a higher and unidirectional activity. In the promoter/reporter gene constructs, transcription initiated from the same positions as in the native gene. Sequence analysis showed that the promoter of the rat AR gene lacks typical TATA and CCAAT box elements, but one SP1 site is located at about 60 bp upstream from the major start site of transcription. Other possible promoter elements are TGTYCT sequences at positions -174 to -179, -434 to -439., -466 to -471, and -500 to -505, resembling half-sites of the glucocorticoid-responsive element (GRE). Furthermore, a homopurine stretch containing a total of 8 GGGGA elements and similar to sequences that are present in several other GC-rich promoters, is located between -89 and -146 bp upstream from the major start site of transcription.
Mol Cell Endocrinol 1990 Nov 12
PMID:The rat androgen receptor gene promoter. 228 81

Multiple forms of human thyroid hormone (T3) receptor have been identified, including true receptors that bind T3 (alpha 1 and beta) and a splicing variant (alpha 2) that does not bind T3. The alpha 1- and beta-receptors activate transcription through interactions with positive thyroid response elements (TREs). The alpha 2 variant is unable to activate transcription and has been reported to inhibit alpha 1 or beta stimulation of positive TREs, a property referred to as dominant negative activity. In this report we have performed studies to assess the functional properties of different members of the thyroid receptor family with regard to both positive and negative transcriptional regulation. The alpha 1-, alpha 2-, and beta-receptors were each coexpressed in JEG-3 cells with either TreTKCAT (CAT = chloramphenicol acetyltransferase), a reporter gene that contains a positive TRE, or TSH alpha CAT, a negatively regulated reporter gene. The alpha 1 and beta isoforms stimulated transcription of TreTKCAT and inhibited TSH alpha CAT transcription in a T3-dependent manner, whereas the alpha 2 variant was inactive. When coexpressed with alpha 1- or beta-receptors, alpha 2 inhibited regulation of positive TREs, but the effects of alpha 2 were modest and only occurred when relatively high doses of receptor were transfected. The alpha 2-receptor variant did not affect negative regulation by alpha 1- or beta-receptors. Thus, in both positive and negative regulation, thyroid hormone receptor isoforms that bind T3 (alpha 1, beta) are functional, whereas the alpha 2 isoform, which does not bind T3, is not functional.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1990 Oct
PMID:Negative and positive transcriptional regulation by thyroid hormone receptor isoforms. 228

We have expressed three forms of human thyroid hormone receptor (hTR alpha 1, alpha 2, and beta) in cultured cells by transient transfection. hTR alpha 1 and beta transfected cells showed increased triiodothyronine (T3) binding capacity, but hTR alpha 2 transfected cells did not. When hTR alpha 1 or beta was cotransfected with pUrGH(S), in which a portion of the rat GH 5' flanking region (-236/-147) was ligated into the CAT reporter plasmid (pUTKAT1), T3 increased CAT gene expression. When hTR alpha 2 was cotransfected with pUrGH(S), T3 did not alter CAT gene expression. When hTR alpha 1 or beta was cotransfected with pUrGH(O), in which a synthetic oligonucleotide representing the TRE from the rat GH 5' flanking region (-189/-160) was substituted for the natural enhancer in pUTKAT1, T3 increased CAT gene expression. When hTR alpha 1 and beta were cotransfected with pUrGH(O), induction by T3 was increased. When hTR alpha 2 was cotransfected with hTR alpha 1 or beta, induction by T3 was decreased. These results indicate that hTR alpha 1 and beta function as native TR, that hTR alpha 1 and beta can recognize the same TRE, that hTR alpha 1 and beta can function additively, and that hTR alpha 2 can inhibit the action of hTR alpha 1 and beta.
Mol Cell Endocrinol 1990 Sep 10
PMID:The roles of three forms of human thyroid hormone receptor in gene regulation. 228 27

A novel cis-acting regulatory element (designated BTE for basic transcription element) was found in the region proximal to the TATA sequence of the P-450c gene by the use of deletion mutations. This DNA element is considered to be involved in the basic transcription of the gene and does not show distinct enhancer activity in itself. Together with the XRE sequence (A. Fujisawa-Sehara, K. Sogawa, M. Yamane, and Y. Fujii-Kuriyama, Nucleic Acids Res. 15:4179-4191, 1987), however, this sequence is required for a high inducible expression of the P-450c gene in response to xenobiotic inducers. The BTE sequence contained the GC box consensus sequence and half of the NF-1-binding consensus or CAT box sequence, but their synthetic oligonucleotides, used as competitors in the gel mobility shift assays, did not compete with the BTE sequence for the binding protein, suggesting that the BTE sequence functions as a different recognition sequence from that for Sp1 or NF-1. Analogous sequences to BTE are found in the region proximal to the TATA sequence of other genes, especially other P-450 genes with different modes of regulation, suggesting that the BTE sequence plays a common regulatory role in basic transcription of genes including a group of the P-450 superfamily. The ubiquitous distribution of nuclear factor(s) binding to this element supports this suggestion.
Mol Cell Biol 1990 Apr
PMID:A novel cis-acting DNA element required for a high level of inducible expression of the rat P-450c gene. 232 4

Previous studies, involving phosphorylation of cytoplasmic proteins and localization of DNA regulatory elements, have suggested that epidermal growth factor (EGF) and 12-O-tetradecanoylphorbol-13-acetate (TPA) have similar actions on prolactin (PRL) gene expression by pituitary (GH) cells. However, little is presently known about whether the actions of these two factors involve common gene-distal intermediates. In the present study, we have employed two approaches to examine this question. Chronic exposure of GH3 cells to TPA, which strongly down-regulates protein kinase C activity, completely inhibited acute TPA stimulation of transient expression of a transfected PRL promoter construct ((-187)PRL-CAT), but did not inhibit EGF stimulation of either accumulation of endogenous PRL mRNA or of expression of (-187)PRL-CAT. Furthermore, the acute stimulatory effects of EGF and TPA on expression of (-187)PRL-CAT were additive. Each of these observations implies that EGF and TPA have gene-distal actions on PRL gene expression that are at least partially non-overlapping.
Mol Cell Endocrinol 1990 Feb 12
PMID:Epidermal growth factor and phorbol ester regulate prolactin gene expression via distinct pathways. 232 87


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