Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The herpes simplex virus thymidine kinase gene (HSV-TK) in combination with ganciclovir (GCV), is currently being used in gene therapy-based clinical trials for cancer treatment. Its therapeutic effect is based on a "bystander effect" whereby HSV-TK gene-modified tumor cells are toxic to nearby unmodified tumor cells when exposed to the antiviral drug GCV. We have recently hypothesized that the in vivo mechanism of this bystander effect is due to alterations in the tumor microenvironment in response to release of cytokines and an infiltration of leukocytes after treatment with HSV-TK gene-modified tumor cells and GCV, which results in tumor regression. Expression of B7, a recently identified costimulatory molecule that is important for T-cell stimulation, has been shown to be modulated by stimulatory cytokines interferon-gamma, tumor necrosis factor-alpha, and inhibited by interleukin-10. In the present study, we investigated whether the cytokines released after HSV-TK and GCV treatment could include the expression of the costimulatory molecules B7-1 and B7-2 and the adhesion molecule (ICAM)-1 in the tumor. Furthermore, we investigated whether this altered environment affected the antitumor properties of host lymphocytes. An in vitro model was developed to establish the effects of HSV-TK gene-modified tumor cells and GCV on tumor infiltrating cells. The murine macrophage cell line (IC21) was exposed to either supernatants or cell lysates collected from a mixture of HSV-TK-transduced (KBALB-STK) and non-transduced (KBALB) murine fibrosarcoma tumor cells previously exposed to GCV (experimental). Immunohistochemical analysis showed a significant expression (P < .0001) of B7-1 and B7-2 post exposure of IC21 cells to either supernatant or lysate. In contrast, the level of expression in IC21 cells exposed to the control lysate or supernatant remained unchanged for B7-1 and B7-2. In vivo analysis for B7-1 and B7-2 expression by immunohistochemistry in tumor tissues from experimental mice receiving HSV-TK gene-modified tumor cells and GCV treatment showed a significant expression of B7.1 (35%, P < .0001) and B7.2 (38.2%, P < .0001) on tumor-infiltrating mononuclear cells. In contrast, tumor-bearing control animals showed low levels of B7-2 expression (5.8%), whereas B7-1 was undetectable, as confirmed by reverse-transcriptase polymerase chain reaction. In addition, a significant up-regulation of ICAM expression (50%) on tumor tissues was observed in the experimental group (P = .0317) as compared with the control group (25%). Furthermore, T cells isolated from experimental mice showed a significant in vitro proliferative response (p = .0202) when exposed to syngeneic tumor cells as compared with the control group. These data demonstrated that the use of HSV-TK gene-modified tumor cells and GCV as a suicide gene in the treatment of an intraperitoneal tumor resulted in the expression of the B7 costimulatory molecules and ICAM-1 adhesion molecule and enhanced proliferative response of host T cells. These findings help to understand the mechanism of tumor cell killing in vivo using HSV-TK gene-modified tumor cells.
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PMID:Expression of costimulatory molecules: B7 and ICAM up-regulation after treatment with a suicide gene. 898 40

The treponemal fla operon is comprised of numerous motility-related genes; however, the initial gene of this operon, tap1, has no known function. A recently developed system to generate specific mutants in Treponema denticola was utilized to determine if Tap1 was essential for motility. T. denticola tap1 and flanking DNA were identified, cloned, and sequenced, and a suicide plasmid that contained tap1 interrupted with an erythromycin resistance cassette (ermF and ermAM) was constructed. Because of potential polar effects from this cassette, a second plasmid that contained tap1 interrupted with a modified erythromycin resistance cassette that lacked the putative ermF transcription terminator was constructed. Electroporation-mediated allelic exchange incorporated the interrupted tap1 genes into the T. denticola chromosome, creating Tap1-deficient mutants. Reverse transcriptase PCR revealed that the erythromycin resistance cassette within tap1 did not terminate fla operon transcription in either mutant. Moreover, the phenotypes of the two mutants were indistinguishable. These mutants lacked motion in liquid culture, were unable to spread on agar plates, and lacked flagellar filaments as determined by electron microscopy. Immunoblots revealed a marked reduction in detectable FlaB flagellar filament protein compared to that of wild type; however, flaB RNA was easily detectable, and transcription levels did not appear to be altered. The basis for the lack of filament protein expression is unknown. Immunoblotting also showed that the flagellar hook protein (FlgE) was synthesized in the Tap1-deficient mutant; however, electron microscopy revealed that the mutant possessed unusual elongated hooks of variable lengths. We propose that treponemal Tap1 is analogous to FliK, which is involved in monitoring the flagellar hook length of Salmonella typhimurium.
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PMID:Insertional inactivation of Treponema denticola tap1 results in a nonmotile mutant with elongated flagellar hooks. 1036 49

We describe here a novel targeting gene therapy strategy to direct gene expression responsive to hepatitis C virus (HCV). The goal was approached by engineering a construct containing the antisense sequence of the transgene and internal ribosome entry site of encephalomyocarditis virus flanked by 5'- and 3'-end sequences of HCV cDNA that contain cis-acting replication elements. Thus, expression of the transgene is only promoted when the minus-strand RNA has been synthesized by the functional replication machinery present in infected cells. Reporter assay and strand-specific reverse transcription-PCR showed selective transgene expression in Huh-7 cells harboring an autonomously replicating HCV subgenome but remaining silent in uninfected cells. Furthermore, using the cytosine deaminase suicide gene as a transgene coupled with recombinant adenovirus delivery, we demonstrated that cytosine deaminase was specifically expressed in replicon cells, resulting in marked chemosensitization of replicon cells to the cytotoxic effects of flucytosine. This new targeting strategy could be extended to other single-stranded RNA viruses encoding the unique RNA-dependent RNA polymerase that has no parallel in mammalian cells.
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PMID:Exploiting cis-acting replication elements to direct hepatitis C virus-dependent transgene expression. 1585 78

Burkholderia cenocepacia, a member of the B. cepacia complex, is an opportunistic pathogen that causes serious infections in patients with cystic fibrosis. We identified a six-gene cluster in chromosome 1 encoding a two-component regulatory system (BCAL2831 and BCAL2830) and an HtrA protease (BCAL2829) hypothesized to play a role in the B. cenocepacia stress response. Reverse transcriptase PCR analysis of these six genes confirmed they are cotranscribed and comprise an operon. Genes in this operon, including htrA, were insertionally inactivated by recombination with a newly created suicide plasmid, pGPOmegaTp. Genetic analyses and complementation studies revealed that HtrA(BCAL2829) was required for growth of B. cenocepacia upon exposure to osmotic stress (NaCl or KCl) and thermal stress (44 degrees C). In addition, replacement of the serine residue in the active site with alanine (S245A) and deletion of the HtrA(BCAL2829) PDZ domains demonstrated that these areas are required for protein function. HtrA(BCAL2829) also localizes to the periplasmic compartment, as shown by Western blot analysis and a colicin V reporter assay. Using the rat agar bead model of chronic lung infection, we also demonstrated that inactivation of the htrA gene is associated with a bacterial survival defect in vivo. Together, our data demonstrate that HtrA(BCAL2829) is a virulence factor in B. cenocepacia.
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PMID:Burkholderia cenocepacia requires a periplasmic HtrA protease for growth under thermal and osmotic stress and for survival in vivo. 1722 Mar 10

Lactic acidosis that is not secondary to tissue hypoperfusion or hypoxemia (type B lactic acidosis) is a rare but potentially fatal condition that has been associated with drugs like metformin, linezolid, and nucleoside reverse-transcriptase inhibitors in patients with HIV. We report the first case of type B lactic acidosis caused by overdose of the serotonin-norepinephrine reuptake inhibitor, venlafaxine. A 55-year-old man with no significant medical history was brought to the emergency department after intentional ingestion of around 80 capsules of venlafaxine (a total dose of over 6000 mg) in an attempt to commit suicide. Complete blood count and comprehensive metabolic panel were unremarkable except for a bicarbonate level of 13 mEq/L and an anion gap of 22 mEq/L. An arterial blood gas revealed a pH of 7.39, partial pressure of CO2 of 19 mm Hg, calculated bicarbonate of 11.5 mEq/L, and a lactate level of 8.6 mmol/L. The patient was started on aggressive intravenous hydration with normal saline along with oral activated charcoal with sorbitol. Repeat laboratory work after 4 hours showed an improvement in anion gap (15 mEq/L) and serum lactate (5.6 mmol/L). The patient remained stable throughout the hospital stay and lactic acidosis resolved in 24 hours. In the absence of hypotension, hypoxemia, kidney or liver dysfunction, myopathy, malignancy, or use of other medications, venlafaxine was the most likely cause of lactic acidosis in our case. Rapid improvement of acidosis was probably related to clearance of the drug.
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PMID:Type B Lactic Acidosis Associated With Venlafaxine Overdose. 2540 96

Genomes of a given bacterial species can show great variation in gene content and thus systematic analysis of the entire gene repertoire, termed the pan-genome, is important for understanding bacterial intra-species diversity, population genetics, and evolution. Here, we analyzed the pan-genome from 30 completely sequenced strains of the human gastric pathogen Helicobacter pylori belonging to various phylogeographic groups, focusing on 991 accessory (not fully conserved) orthologous groups (OGs). We developed a method to evaluate the mobility of genes within a genome, using the gene order in the syntenically conserved regions as a reference, and classified the 991 accessory OGs into five classes: Core, Stable, Intermediate, Mobile, and Unique. Phylogenetic networks based on the gene content of Core and Stable classes are highly congruent with that created from the concatenated alignment of fully conserved core genes, in contrast to those of Intermediate and Mobile classes, which show quite different topologies. By clustering the accessory OGs on the basis of phylogenetic pattern similarity and chromosomal proximity, we identified 60 co-occurring gene clusters (CGCs). In addition to known genomic islands, including cag pathogenicity island, bacteriophages, and integrating conjugative elements, we identified some novel ones. One island encodes TerY-phosphorylation triad, which includes the eukaryote-type protein kinase/phosphatase gene pair, and components of type VII secretion system. Another one contains a reverse-transcriptase homolog, which may be involved in the defense against phage infection through altruistic suicide. Many of the CGCs contained restriction-modification (RM) genes. Different RM systems sometimes occupied the same (orthologous) locus in the strains. We anticipate that our method will facilitate pan-genome studies in general and help identify novel genomic islands in various bacterial species.
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PMID:A Novel Approach to Helicobacter pylori Pan-Genome Analysis for Identification of Genomic Islands. 2750 80