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

---

Query: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In herpes simplex virus (HSV)-infected cells, the transcription of immediate-early (alpha) genes is regulated by a virion component, the alpha gene trans-inducing factor (alpha TIF). This protein forms a complex with cellular factors and TAATGARAT motifs present in one or more copies in the promoters of all alpha genes. We have characterized the bovine herpesvirus 1 (BHV-1) homolog of this protein. Like its HSV counterpart, the BHV alpha TIF was synthesized in the later stages of infection and could be demonstrated to be a component of purified virions. In transient expression assays, BHV alpha TIF was a strong transactivator and stimulated the activity of IE-1, the major BHV-1 alpha gene promoter, with an efficiency comparable to that of HSV alpha TIF. This stimulation was largely dependent on a TAATGAGCT sequence present in a single copy in IE-1, and BHV alpha TIF, in conjunction with cellular factors, formed a complex with oligonucleotides containing this sequence. Despite these similarities between the two alpha TIFs, our preliminary observations suggest that the proteins may activate transcription by different mechanisms. Although BHV alpha TIF strongly transactivated IE-1, it differed from its HSV counterpart in that the carboxyl terminus of BHV alpha TIF, when fused to the DNA-binding domain of GAL4, was a relatively poor stimulator of a promoter containing GAL4-binding sites. Also unlike HSV alpha TIF, removal of the carboxyl terminus of BHV alpha TIF reduced but did not eliminate the ability of the protein to transactivate IE-1. These results are discussed in view of the structural similarities and differences among the alpha TIFs of alphaherpes-viruses.
...
PMID:Protein and DNA elements involved in transactivation of the promoter of the bovine herpesvirus (BHV) 1 IE-1 transcription unit by the BHV alpha gene trans-inducing factor. 803 88

In contrast to previous findings, the Ab4 isolate of equid herpesvirus-1 (EHV-1) was shown to share homology with the G9 isolate of equid herpesvirus-2 (EHV-2). Using Southern blotting and stringent hybridization conditions, a significant proportion of this cross-hybridization was identified by the immediate-early gene-3 (IE-3) probe from herpes simplex virus-1 (HSV-1). The HSV-1 UL48 gene probe (encoding the IE gene transactivating protein VmW65, which is also known as alpha-TIF or VP16) was used to identify and isolate its counterpart in EHV-1. The relevance of shared homology to transactivation is being investigated.
...
PMID:Cross-hybridization of equid herpesvirus-2 (EHV-2) and herpes simplex virus-1 (HSV-1) genes to equid herpesvirus-1 (EHV-1). 810 47

The 65-kDa herpes simplex virus type 1 encoded alpha trans-induction factor (alpha TIF or VP16) has two important functions: it is required for the efficient transcriptional induction of the alpha or immediate-early genes, and it acts as an essential structural component of the virion. The transcription properties of alpha TIF have been well studied in vitro. The protein is a powerful inducer of RNA polymerase II-directed transcription and, similar to the cellular transcriptional transactivators GAL4 and CGN4, contains separable DNA binding and transactivation domains. In contrast, little is known about the structural function of alpha TIF because this function can be studied only during virus replication and structural mutants are lethal. The in vivo analysis of alpha TIF is further complicated by the likelihood that the transcription and structural functions are not entirely separable. In this study, we take an alternate approach toward the development of alpha TIF mutants and their subsequent characterization. Rather than analyzing the effects of intragenic mutations, we have examined the properties of a mutant virus which expresses an alpha TIF fusion protein containing 61 amino acids of another herpes simplex virus type 1 virion protein, VP13/14, fused to its C terminus. This is the first report which demonstrates that the C-terminus of alpha TIF can tolerate the addition of an adjacent protein domain without compromising its transactivation function in vivo. Moreover, the VP13/14 sequences do not interfere with the protein-protein interactions required for virion targeting and assembly.
...
PMID:An 85-kilodalton herpes simplex virus type 1 alpha trans-induction factor (VP16)-VP13/14 fusion protein retains the transactivation and structural properties of the wild-type molecule during virus infection. 810 36

Mouse LTK- cells were transfected with a pair of defective Herpes simplex virus thymidine kinase (tk) genes. One tk gene had an 8-bp insertion mutation while the second gene had a 100-bp inversion. Extrachromosomal homologous recombination leading to the reconstruction of a functional tk gene was monitored by selecting for tk positive cells using medium supplemented with hypoxanthine/aminopterin/thymidine. To assess whether the search for homology may be a rate-limiting step of recombination, we asked whether the presence of an excess number of copies of a tk gene possessing both the insertion and inversion mutations could inhibit recombination between the singly mutated tk genes. Effective competitive inhibition would require that homology searching (homologous pairing) occur rapidly and efficiently. We cotransfected plasmid constructs containing the singly mutated genes in the presence or absence of competitor sequences in various combinations of linear or circular forms. We observed effective inhibition by the competitor DNA in six of the seven combinations studied. A lack of inhibition was observed only when the insertion mutant gene was cleaved within the insertion mutation and cotransfected with the two other molecules in circular form. Additional experiments suggested that homologous interactions between two DNA sequences may compete in trans with recombination between two other sequences. We conclude that homology searching is not a rate-limiting step of extrachromosomal recombination in mammalian cells. Additionally, we speculate that a limiting factor is involved in a recombination step following homologous pairing and has a high affinity for DNA termini.
...
PMID:The search for homology does not limit the rate of extrachromosomal homologous recombination in mammalian cells. 815 Feb 86

The equine herpesvirus 1 (EHV-1) homolog of herpes simplex virus type 1 ICP22 is differently expressed from the fourth open reading frame of the inverted repeat (IR4) as a 1.4-kb early mRNA and a 1.7-kb late mRNA which are 3' coterminal (V. R. Holden, R. R. Yalamanchili, R. N. Harty, and D. J. O'Callaghan, J. Virol. 66:664-673, 1992). To extend the characterization of IR4 at the protein level, the synthesis and intracellular localization of the IR4 protein were investigated. Antiserum raised against either a synthetic peptide corresponding to amino acids 270 to 286 or against a TrpE-IR4 fusion protein (IR4 residues 13 to 150) was used to identify the IR4 protein. Western immunoblot analysis revealed that IR4 is expressed abundantly from an open reading frame composed of 293 codons as a family of proteins that migrate between 42 to 47 kDa. The intracellular localization of IR4 was examined by cell fractionation, indirect immunofluorescence, and laser-scanning confocal microscopy. These studies revealed that IR4 is localized predominantly in the nucleus and is dispersed uniformly throughout the nucleus. Interestingly, when IR4 is expressed transiently in COS-1 or LTK- cells, a punctate staining pattern within the nucleus is observed by indirect immunofluorescence. Cells transfected with an IR4 mutant construct that encodes a C-terminal truncated (19 amino acids) IR4 protein exhibited greatly reduced intranuclear accumulation of the IR4 protein, indicating that this domain possesses an important intranuclear localization signal. Western blot analysis of EHV-1 virion proteins revealed that IR4 proteins are structural components of the virions. Surprisingly, the 42-kDa species, which is the least abundant and the least modified form of the IR4 protein family in infected cell extracts, was the most abundant IR4 protein present in purified virions.
...
PMID:Identification and characterization of the ICP22 protein of equine herpesvirus 1. 820 8

Transient expression assays measuring induction of an equine herpesvirus 1 (EHV-1) immediate early (IE) promoter construct, pIE beta gal, were used to examine trans-induction of the IE gene (IR1) promoter by superinfection with intact EHV-1 (Kentucky A strain), uv-inactivated EHV-1, or EHV-1 stocks highly enriched for defective interfering particles (DIPs), and by cotransfection with plasmids containing EHV-1 DNA fragments. Our results confirm reports by others in that the IE promoter can be induced by both intact and uv-inactivated EHV-1 and provide evidence that DIP-rich virus stocks also possess transinducing activity. Furthermore, IE promoter activity was induced by cotransfection of the promoter construct with either of two plasmids containing restriction enzyme DNA fragments that span ORF12, the putative EHV-1 homolog of the herpes simplex virus 1 (HSV-1) alpha-trans-inducing factor (alpha-TIF) gene, UL48, or by cotransfection with an isolated DNA fragment containing only ORF12, while no transactivation was associated with plasmids linearized by restriction enzyme digestion at a single site within ORF12. These data indicate that, despite its predicted lack of an acidic carboxy terminus (a region essential in HSV-1 alpha-TIF for trans-inducing activity), the EHV-1 ORF12 product is a functional alpha-TIF.
...
PMID:Transcriptional control of the equine herpesvirus 1 immediate early gene. 824 1

Analysis of the promoter for the herpes simplex virus (HSV) immediate-early (alpha) gene alpha 0 in a short-term transient expression assay revealed that a SacI-to-NcoI fragment from -786 to +148 relative to the cap site directed the synthesis of chloramphenicol acetyltransferase when the fragment was present in either orientation. Although the constitutive levels of promoter activity were similar with either orientation, the reverse-orientation promoter was not induced in response to infection with HSV. Analysis of sequences composing the putative promoter in the opposite orientation revealed the presence of important regulatory elements associated with alpha promoters. These include an alpha-trans-inducing factor (alpha-TIF)-like response element, a high-affinity ICP4-binding site, numerous Sp1-binding sites, and a TATA box. Sequences contained within this region formed specific DNA-protein complexes in extracts from mock-infected and HSV-infected HeLa cells. Transient expression assays revealed that this sequence was positively regulated by the alpha 0 and alpha-TIF genes but negatively regulated by alpha 4. Finally, nuclear run-on transcription assays revealed that this promoter is active in its correct genomic context during the course of virus infection. We suggest that the promoter is a hybrid between an alpha and beta promoter because it exhibits maximal expression at 8 h postinfection and is expressed in the presence of cycloheximide.
...
PMID:Identification of a promoter mapping within the reiterated sequences that flank the herpes simplex virus type 1 UL region. 838 Apr 59

The gene products of herpes simplex virus type 1 UL46 and UL47 enhance the efficiency of alpha TIF (VP16)-mediated alpha gene expression through an unknown mechanism of action. To further characterize the function of the UL46- and UL47-encoded proteins during virus infection, a series of isogenic herpes simplex virus type 1 strain F-derived UL46 and UL47 single-deletion mutants and a UL46/47 double-deletion mutant were constructed and compared with the wild type. Analysis of purified virions obtained from the UL46 deletion mutant showed for the first time that UL46 encoded the viron tegument phosphoproteins VP11 and VP12 (VP11/12). Similar analyses of the UL47 deletion mutants confirmed an earlier report by McLean et al. that UL47 also encoded two virion tegument phosphoproteins, VP13 and VP14 (VP13/14) (G. McLean, F. Rixon, N. Langeland, L. Haarr, and H. Marsden, J. Gen. Virol. 71:2953-2960, 1990). Kinetic analysis demonstrated a delay of approximately 2 h in the appearance of thymidine kinase (TK) activity in all of the UL46 and UL47 single-deletion mutants. In the UL46/47 double-deletion mutant, the delay in TK activity increased twofold, suggesting that the proteins encoded by UL46 and UL47 may act at the same level. Since the delay in TK expression occurred within the first 4 h of infection, the actions of VP11/12 and VP13/14 resulted from their virion association and not from their de novo synthesis as late (beta gamma and gamma) genes. Densitometric analysis of purified virions showed that the levels of VP11/12 and VP13/14 in the virion tegument were near the molar ratios of alpha TIF. On the basis of these observations, we predict that the abilities of UL46 and UL47 to enhance alpha TIF-mediated transcription could result from a stoichiometric association of VP11/12 and VP13/14 with alpha TIF within the infecting virion.
...
PMID:Herpes simplex virus type 1 UL46 and UL47 deletion mutants lack VP11 and VP12 or VP13 and VP14, respectively, and exhibit altered viral thymidine kinase expression. 838 6

The assembly of specific multiprotein complexes on the herpes simplex virus alpha/IE (immediate early) enhancer elements requires the interactions of the Oct-1 POU homeodomain, the viral alpha TIF (alpha-trans-induction factor) (VP16), and at least one additional cellular factor, the C1 factor. The C1 factor interacts directly with alpha TIF, likely forming an intermediate protein complex that recognizes the Oct-1 homeodomain-DNA complex. The biochemical purification of the mammalian C1 factor suggests that it is composed of multiple subunits of related, but heterogeneous, polypeptides. The interaction of a subset of these polypeptides with alpha TIF is stimulated by post-translational modifications of the C1 proteins, suggesting that this factor may be a critical target for the regulation of the herpes simplex virus alpha/IE transcription.
...
PMID:Purification of the cellular C1 factor required for the stable recognition of the Oct-1 homeodomain by the herpes simplex virus alpha-trans-induction factor (VP16). 845 22

Imaging the expression of successful gene transduction has been demonstrated in vivo for the first time by using an appropriate combination of "marker gene" and "marker substrate" in an experimental animal model. The herpes simplex virus 1 thymidine kinase (HSV1-tk) gene was selected as an example of a marker gene, and the recombinant STK retrovirus containing HSV1-tk was used to transduce RG2 glioma cells in vitro and in vivo. RG2TK+ cell lines expressing the HSV1-tk gene and three potential marker substrates for the HSV1-TK enzyme were evaluated. Radiolabeled 5-iodo-2'-fluoro-2'deoxy-1-beta-D-arabinofuranosyluracil (FIAU) was shown to be a substantially better marker substrate for the HSV1-TK enzyme than 5-iodo-2'-deoxyuridine or ganciclovir. The magnitude of FIAU accumulation in different RG2TK+ clones corresponded to their sensitivity to ganciclovir and to the level of HSV1-tk mRNA expression. Imaging the expression of HSV1-tk in transduced RG2 tumor cells was demonstrated in animals using quantitative autoradiography; 2-[14C]FIAU accumulation was shown to be high in RG2TK+ brain tumors growing in one hemisphere and very low in nontransduced RG2 tumors in the contralateral hemisphere. Transduction of RG2 tumor cells with the HSV-tk gene in vivo resulted in tumors which accumulated FIAU to high levels and produced clearly defined images. Given the level of FIAU accumulation in the transduced tumors, it is likely that a clinically applicable method for imaging HSV1-tk gene expression can be implemented using existing clinical imaging techniques.
...
PMID:Imaging the expression of transfected genes in vivo. 852 3


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---