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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We find that tamoxifen is a potent activator of estrogen receptor (ER)- mediated induction of promoters regulated by AP-1 sites including the human collagenase gene promoter and constructs in which an AP-1 site is fused to the herpes thymidine kinase promoter. This contrasts with the inability of tamoxifen to activate otherwise identical promoters bearing classical estrogen response elements. Tamoxifen agonism at AP-1 sites is cell type specific, occurring in cell lines of uterine, but not of breast, origin. It thus parallels tamoxifen agonism in vivo. AP-1 proteins such as Jun or Jun/Fos are needed for tamoxifen stimulation, and tamoxifen increases the transcriptional efficiency of these proteins even when they are provided at optimal amounts. The DNA binding domain (DBD) of ER is required for tamoxifen activation at AP-1 sites. In contrast, estrogen activation is partially independent of this domain. This suggests the existence of two pathways of ER action at AP-1: an alpha (DBD-dependent) pathway activated by tamoxifen, and a beta (DBD-independent) pathway activated by estrogen. Fusing VP16 transcriptional activation functions to ER potentiates the beta, but not the alpha, pathway. We discuss models for the two pathways and the possibility that the AP-1 pathway is a major route by which ER affects target tissue growth and differentiation in vivo.
Mol Endocrinol 1995 Apr
PMID:Tamoxifen activation of the estrogen receptor/AP-1 pathway: potential origin for the cell-specific estrogen-like effects of antiestrogens. 765 88

The 5'-flanking region of the gene for Pit-1, a pituitary-specific transcription factor, was isolated from a rat liver genomic library and sequenced. Expression of a reporter construct containing Pit-1 promoter sequences linked to the bacterial chloramphenicol acetyltransferase (CAT) gene was assessed by transient transfection in rat pituitary GH4C1 cells. Treatment of transfected cells with either dexamethasone (DEX) for 48 h or the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) for the final 20 h of the 48-h posttransfection period had minimal effects on CAT expression. However, CAT activity was elevated about 20-fold when transfected cells were treated with both DEX and TPA. This apparent synergistic activation was lost when DEX treatment was also limited to the final 20 h of the 48-h posttransfection period, suggesting that a time-dependent accumulation of a DEX-induced gene product might be involved. This putative DEX-induced product appeared to be relatively stable, because synergistic activation was observed in cells treated with DEX alone for 36 h, followed by a 10-h incubation without DEX before the addition of TPA. The Pit-1 gene promoter region between -210 and -142 from the transcription start site conferred synergistic regulation by DEX and TPA when placed upstream of position -105 in the herpes viral thymidine kinase promoter.(ABSTRACT TRUNCATED AT 250 WORDS)
Mol Endocrinol 1994 Oct
PMID:A sequence in the rat Pit-1 gene promoter confers synergistic activation by glucocorticoids and protein kinase-C. 785 49

9-(2-Phosphonylmethoxyethyl)adenine (PMEA) is a new antiviral agent with activity against herpes viruses and retroviruses, including human immunodeficiency virus, but its metabolism and mechanism of action remain unclear. We have isolated a human T lymphoid cell line (CEMr-1) that is resistant to the antiproliferative effects of PMEA. The antiviral effects of PMEA against human immunodeficiency virus-1 infection were also greatly reduced in CEMr-1 cells, compared with the parental cells. This mutant showed cross-resistance to the related acyclic nucleoside phosphonates 9-(2-phosphonylmethoxyethyl)diaminopurine and 9-(2-phosphonylmethoxyethyl)guanine and the lipophilic prodrug bis(pivaloyloxymethyl)-9-(2-phosphonylmethoxyethyl)adenine-( bispome-PMEA), as well as partial resistance to the purine nucleosides 2-chlorodeoxyadenosine, 2-fluro-9-beta-D-arabinosylfuranosyladenine, and adenosine, but did not show resistance to 2'-deoxyadenosine or 9-beta-D-arabinosylfuranosyladenine. We compared the uptake and metabolism of [3H]PMEA and [3H]-bispom-PMEA in the mutant and parental cells. The analysis of radioactive products by high pressure liquid chromatography revealed marked alterations in the ability of the mutant cell line to accumulate PMEA and its anabolites, compared with the parental cells. Accumulation of PMEA, PMEA monophosphate, and PMEA bisphosphate (major metabolites formed with either PMEA or bispom-PMEA) decreased by 50, 95, and 97%, respectively. Compared with the parental cells, the variant cells showed a approximately 7-fold increase in the rate of efflux of PMEA and a 2-fold decrease in the activity of adenylate kinase. In contrast, other enzymes of nucleotide metabolism, such as adenosine kinase, deoxycytidine kinase, and 5-phosphoribosyl-1-pyrophosphate synthetase, showed no significant change in the two cell lines. Overall, these results suggest that the mutation in this resistant cell line is of a novel type, involving an alteration in the cellular efflux of PMEA as the major basis for the resistant phenotype.
Mol Pharmacol 1995 Feb
PMID:A human T lymphoid cell variant resistant to the acyclic nucleoside phosphonate 9-(2-phosphonylmethoxyethyl)adenine shows a unique combination of a phosphorylation defect and increased efflux of the agent. 787 49

A rapid and sensitive one-step polymerase chain reaction (PCR) assay was developed for use in identifying type I and type II herpes simplex virus (HSV). Although the nucleotide sequences of the two HSV subtypes are quite similar, common and type-specific sequences 20 nucleotides in length could be deduced in the thymidine kinase gene. Oligonucleotide primers targeted to the type-specific regions generated products of different sizes that served to distinguish two HSV types. Type-specific PCR amplification products were verified by restriction enzyme digestion. Specificity of the HSV PCR was established by the lack of amplification of other herpes-group viruses including cytomegalovirus, Epstein-Barr virus, and varicella zoster virus. Extraction of DNA from clinical materials (throat swabs, vesicular swabs, cerebrospinal fluid and eye discharge) yielded an amplification product of the predicted size for each HSV type. Thus, this PCR system provides a rapid, sensitive and specific assay that can supplement the currently available modalities for detecting and typing HSV.
Mol Cell Probes 1994 Jun
PMID:One-step determination of herpes simplex virus types I and II by polymerase chain reaction. 796 91

1. Herpesvirus infection with genetically engineered vectors is a way to deliver foreign gene products to various cell populations in culture and in vivo. Selective neuronal gene expression can be achieved using the neuron-specific enolase (NSE) promoter regulating expression of a transgene placed in and delivered by a herpesvirus vector. 2. We sought to determine the anatomical specificity and efficiency of herpesvirus-mediated gene transfer into the rat brain following placement of virus particles carrying a transgene (lacZ) under control of the NSE promoter. The virus utilized was thymidine kinase (TK) deficient and therefore replication deficient in the brain. 3. Infusion of 10(6) plaque-forming units of virus into the striatum caused a limited number of striatal neurons to express the lacZ transgene mRNA and protein product 7 days postinfection. In addition, small numbers of neurons expressing the transgene mRNA and protein were found ipsilateral to the viral injection in the frontal cortex, substantia nigra pars compacta, and thalamus. Neurons at these anatomic loci project directly to the striatal injection site. No other cells within the brains of injected animals expressed the lacZ gene. 4. While this herpesvirus NSE vector was capable of introducing novel functional genetic information into postmitotic neurons within defined neuroanatomic constraints, the numbers of neurons expressing detectable levels of beta-galactosidase was minimal. The calculated efficiency of delivery and transgene expression at 7 days postinfection was 1 transgenic neuron per 10(4) virus particles infused. 5. We conclude that NSE probably is not an optimal promoter for use in gene delivery to CNS neurons in herpesvirus vectors and that the efficacy of gene delivery using other neuron-specific promoters placed at various sites in the herpes viral genome needs to be explored.
Cell Mol Neurobiol 1993 Oct
PMID:Herpesvirus-mediated gene delivery into the rat brain: specificity and efficiency of the neuron-specific enolase promoter. 811 22

A recently identified sequence motif, referred to as "C3HC4" (also "RING finger" and "A Box") for its distinctive pattern of putative metal-binding residues, has been found in a wide range of proteins. In a previous paper we described the expression and purification of fragments encompassing this motif from the Vmw110 (IPC0) protein family. We showed that the equine herpes virus protein binds zinc ions and adopts a beta beta alpha beta fold. We now report the tertiary structure of this domain in solution, as determined by two-dimensional 1H-NMR An amphipathic alpha-helix lies along one surface of a triple-stranded beta-sheet. Four pairs of metal-binding residues sequester two zincs at distinct tetrahedral sites. The first and third pairs bind one metal ion, while the second and fourth pairs bind the other, forming an interleaved whole. The first and the fourth pairs are contained within two prominent, well-defined loops related by an approximate dyad symmetry. Conserved residues within the helix, sheet and loops contribute to a compact hydrophobic core. The region comprising the first two beta-strands and the alpha-helix has remarkable structural similarity with a TFIIIA type of zinc finger, even though the C3HC4 domain appears not to bind specifically to DNA or RNA. Using site-directed mutagenesis we demonstrate that exposed polar side-chains of the C3HC4 alpha-helix are essential for trans-activation of gene expression by an intact herpes virus regulatory protein.
J Mol Biol 1994 Mar 25
PMID:Structure of the C3HC4 domain by 1H-nuclear magnetic resonance spectroscopy. A new structural class of zinc-finger. 812 34

Cytomegalovirus (CMV) is a member of the herpes virus group. Infection results in a variety of disorders which depend largely on the immune status of the host. A well known property of CMV is that after primary infection the virus persists in the body of the host resulting in latency. Severe immunodepression or immunodeficiency can cause reactivation of the virus from its latent state, leading to endogenous reinfection. In contrast to other herpes viruses, such as herpes simplex virus which persists in neurons, and Epstein Barr virus which persists in B lymphocytes, little is known about the localization of latent CMV. In order to obtain more insight in the organ or cell type serving as a reservoir for latent CMV, it is important to know more about the course of natural infection and the cells and organs involved. When more information is available about the localization of latent virus, studies concerning the physical state of viral DNA or the extent of viral transcription and/or translation will follow in the near future. In this review some properties of the epidemiology and transmission of human CMV, as well as data about acute infection will be given. In addition, some characteristics of the localization of latent CMV and the physical state of the virus will be discussed. Where necessary, particularly regarding insight into CMV-host interactions, knowledge of animal, particularly murine, rat and guinea pig CMV infections, will be discussed.
Virchows Arch B Cell Pathol Incl Mol Pathol 1993
PMID:Cytomegalovirus and latency: an overview. 814 53

A highly conserved, cysteine-rich region plays a crucial role in the function of a family of regulatory proteins encoded by alpha herpes viruses. The so-called C3HC4 motif spans approximately 60 residues and has been predicted to bind zinc. This motif occurs in a number of other viral and cellular proteins, many of which appear to be involved in some aspect of the regulation of gene expression. We have cloned and expressed in bacteria a portion of immediate-early protein Vmw110 of herpes simplex virus type 1 that encompasses the C3HC4 motif, and the equivalent regions from the homologous proteins of varicella zoster virus and equine herpes virus type 1 (EHV-1). All three polypeptides were purified and found to bind zinc stably. None of the three interacted significantly with either DNA or RNA under our assay conditions. The EHV-1 domain yielded interpretable proton nuclear magnetic resonance spectra. Assignment of resonances and analysis of nuclear Overhauser effects revealed its secondary structure. Starting from the N terminus, this consists of an ordered but irregular loop, the first two strands of a triple-stranded antiparallel beta-sheet, two turns of an alpha-helix, a second irregular loop, and the third strand of the beta-sheet. It appears that, taking the cysteine and histidine residues in turn, cysteine residues I, II, IV and V co-ordinate one zinc atom while the histidine residue and cysteine residues III, VI and VII co-ordinate a second zinc atom. This arrangement of secondary structure differs from that found in other characterized zinc-containing proteins.
J Mol Biol 1993 Dec 20
PMID:A novel arrangement of zinc-binding residues and secondary structure in the C3HC4 motif of an alpha herpes virus protein family. 826 11

CdG, the carbocyclic analog of 2'-deoxyguanosine, is active against herpes, hepatitis B, and human cytomegaloviruses. We have studied the interaction of the tritiated enantiomers of CdG with the herpes simplex virus type 1-specific thymidine kinase (HSV-1 TK) and have examined their metabolism in uninfected and HSV-1-infected cells. D- and L-CdG were equally effective competitive inhibitors of the phosphorylation of thymidine (dThd) by the partially purified HSV-1 TK (Ki values were 2.1 and 3.4 microM, respectively) and were also equal as substrates (Km values were 17 and 26 microM, respectively, and Vmax values of the enantiomers were equal and about 50% greater than the Vmax for dThd). The partially purified enzyme preparation, which contained cellular nucleotide kinase activities (pyruvate kinase also was present in the assay medium), converted D-CdG almost exclusively to the triphosphate and L-CdG almost exclusively to the monophosphate. Similarly, in virus-infected cells the D-enantiomer was converted predominantly to the triphosphate and the L-enantiomer predominantly to the monophosphate. In uninfected cells the results were qualitatively similar. In CEM cells deoxycytidine (dCyd) kinase (EC 2.7.1.74) seemed to be the enzyme principally responsible for the phosphorylation of both enantiomers, as shown by competition studies. Thus, both the HSV-1 TK and cellular dCyd kinase (of CEM cells) showed no selectivity for the enantiomers of CdG. This lack of enantiomeric specificity has obvious implications for the design of inhibitors of both viral proliferation and cellular metabolism.
Mol Pharmacol 1993 Dec
PMID:Phosphorylation of the enantiomers of the carbocyclic analog of 2'-deoxyguanosine in cells infected with herpes simplex virus type 1 and in uninfected cells. Lack of enantiomeric selectivity with the viral thymidine kinase. 826 63

Double polymerase chain reaction (PCR) assays with nested primers have been applied in a routine laboratory for the diagnosis of herpes-, pesti- and retroviral infections of animals. Various methods and tools have been tested to prevent and to eliminate false positive results as well as to visualize the PCR products (amplicons). The u.v. and DNase treatments proved to be unsuitable for decontamination of PCR mixtures contaminated with amplicons shorter than 380 bp. By constructing special tube-holders and openers, and by applying a simple technique of pipetting, the false-positive PCR results were eliminated. The PCR products were visualized by three simple methods. The solid phase colorimetric method termed 'Detect Immobilized Amplified DNA' (DIANA) has been adapted to microplate. The other method, termed 'Colorimetric Detection Assay on Filter' (CODAF), proved to be very rapid. However, despite these advantages of DIANA and CODAF, henceforward the nucleic acid hybridization methods were found most reliable for safe identification of PCR amplicons. In order to simplify the hybridization, various non-radioactive labelling methods of oligonucleotide probes were compared. Biotinylation at the 5' end by means of oligonucleotide synthesis was the most simple and practical labelling method in this laboratory. The routine applicability of hybridization was further simplified by constructing a robot device, which automatically performs filter-hybridization and subsequently develops the signals derived from the biotinylated hybrids.
Mol Cell Probes 1993 Jun
PMID:Experiences on the application of the polymerase chain reaction in a diagnostic laboratory. 836 70


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