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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Specific binding of radiolabelled FSH and LH to rat ovaries was demonstrated at the age of 7 days. However, when the biological response to LH and FSH was monitored by cAMP production in vitro, the FSH response appeared earlier than that of LH, on days 4 and 7, respectively. Cholera toxin stimulated cAMP production even in fetal ovaries, suggesting the presence of functional post-receptor machinery of cAMP production. Hence, the appearance of the functional gonadotropin receptor probably plays a key role in the onset of postnatal ovarian steroidogenesis. To test the effect of gonadotropin suppression during postnatal ovarian development, a potent GnRH antagonist was administered to neonatal animals between days 1-6 or 1-9 of life. The ovarian responsiveness to FSH developed even in the absence of normal gonadotropin levels, but that to LH was suppressed after the longer antagonist treatment. The temporal relationship between the onset of LHR gene expression, i.e. transcription, and translation to functional receptor protein was thereafter investigated using the reverse transcriptase-polymerase chain reaction (RT-PCR) technique. The measurements revealed the existence only of truncated versions of LHR mRNA in the fetal ovary from day 17 of gestation up to day 7 of postnatal life. With the onset of the receptor function around day 7, also larger mRNA transcripts, corresponding to the full-length receptor protein appeared. Our findings suggest that the LHR gene may be constitutively expressed in the ovary and a change in the alternative splicing pattern may cause the onset of translation of a functional receptor protein.
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PMID:Ontogeny of gonadotropin action in the rat ovary. 134 71

A rapid method for the detection of hog cholera virus (HCV) in infected tissues, using polymerase chain reaction (PCR) was developed. Total RNA isolated from HCV-infected tissues was reverse transcribed with AMV reverse transcriptase and the resulting complementary DNA was amplified by Taq DNA polymerase in the presence of two HCV-specific primers. The amplified DNA fragment was detected by agarose gel electrophoresis. The sensitivity of this method was at 10(4) TCID50 of HCV. The sensitivity increased approximately 1000-fold when the DNA was reamplified with a set of nested primers. DNA sequencing analysis of the PCR products revealed that the HCV sequence amplified from a local field isolate was highly homologous to the HCV Alfort strain. This method may be useful for pathological and epidemiological studies of HCV in pigs.
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PMID:Rapid detection of hog cholera virus in tissues by the polymerase chain reaction. 181 55

Currently available evidence supports the hypothesis that insulin-like growth factor-I (IGF-I) secreted by small preantral follicles may be involved in stimulating the initial differentiation of the theca interna and, in particular, expression of the LH receptor in pre-theca cells. To test this hypothesis, we examined the effects of IGF-I on LH receptor mRNA expression in theca-interstitial cells (TIC) isolated from the ovaries of hypophysectomized immature rats by percoll gradient centrifugation. TIC (3.5 x 10(4) viable cells/well) were cultured up to 6 days with and without LH (0-10 ng/ml) and IGF-I (0-100 ng/ml). Androsterone in the medium was measured by RIA, and LH receptor mRNA was measured by specific reverse transcriptase-polymerase chain reaction assay. LH receptor mRNA was low in control (untreated) TIC. IGF-I stimulated a dose-related increase (2-fold) in LH receptor mRNA at 2 days (ED50 = 9.0 +/- 1.9 ng/ml) that remained constant at 4 days and then declined to basal levels at 6 days. LH stimulated a dose-related (ED50 = 17.6 +/- 1.0 pg/ml) increase in LH receptor mRNA that reached a maximum of 4-fold at 2 days. At 4 days, LH down-regulated LH receptor mRNA below basal levels, and it had no effect at 6 days. Addition of IGF-I (30 ng/ml) to LH-treated TIC abolished the stimulatory effect of LH throughout the culture period. LH receptor mRNA was highly sensitive to LH since the ED50 was approximately 2.5-fold lower than for stimulation of androsterone production (39.8 +/- 3.8 pg/ml). To understand the molecular mechanism of the synergistic stimulation of androgen production by IGF-I and LH, the effects of IGF-I on the cAMP/protein kinase A (PKA) signaling pathway were examined. When freshly isolated TIC were challenged with IGF-I alone (30 ng/ml), there was no effect on cAMP production or PKA activity, but IGF-I augmented LH stimulation of cAMP production slightly at high concentrations of LH and blocked stimulation of PKA activity by a saturating concentration of LH (3 ng/ml), suggesting that IGF-I increased LH down-regulation of PKA. We next examined the effects of IGF-I on LH receptor number. When TIC were placed into culture, LH/hCG binding sites decreased to approximately 35% of the initial number at 24 h and 25% at 2 days. This decrease was accompanied by a similar loss of cholera toxin- and hCG-stimulated cAMP production.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Insulin-like growth factor-I regulation of luteinizing hormone (LH) receptor messenger ribonucleic acid expression and LH-stimulated signal transduction in rat ovarian theca-interstitial cells. 752 76

The protein E-specific lysis mechanism of the Escherichia coli-specific bacteriophage PhiX174 was employed to produce Vibrio cholerae ghosts (VCG). VCG consist of both rounded and collapsed cells that have lost their cytoplasmic contents through an E-specific hole in the cell envelope. These ghosts are proposed as non-living material for immunization against cholera. A specific membrane anchor sequence was used to insert the human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) fusion protein into the cell envelope of V. cholerae. The identity of the expression products was confirmed by Western blot analysis employing an RT-specific monoclonal antibody. HIV-1 RT was chosen as a model for the purpose of evaluating heterologous gene expression in V. cholerae and the carrier potential of VCG. Intraperitoneal immunization of mice was used to evaluate the immunogenic potential of VCG. Preliminary results showed significant seroconversions to intact whole-cell vibrio antigens in mice immunized with VCG or a heat-killed whole-cell vibrio preparation.
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PMID:Production of Vibrio cholerae ghosts (VCG) by expression of a cloned phage lysis gene: potential for vaccine development. 753 Aug 88

Despite pathophysiologic effects including diarrhea, cholera toxin (CT) is a potent mucosal immunogen and adjuvant. We investigated the influence of CT on T helper (Th)-type 1 (Th1) and Th2 cell-regulated Ag-specific B cell isotype and IgG subclass Ab responses elicited when the toxin was co-administered orally with different protein Ags. When mice were orally immunized with tetanus toxoid (TT) and CT as adjuvant, this regimen induced TT-specific secretory IgA responses in the gastrointestinal tract as well as serum IgG, including IgG1 and IgG2b subclasses, and IgA responses. This oral regimen also induced TT- and CT-B-specific IgE responses. In addition, CT also elicited adjuvant effects for Ag-specific IgG1, IgE, and IgA responses when two other protein Ags, OVA and hen egg white lysozyme, were given by the oral route. Quantitative reverse transcriptase-PCR was performed to assess levels of mRNA for Th1 (IFN-gamma) and Th2 (IL-4) cytokine expression in TT-stimulated CD4+ T cell cultures. Both Peyer's patches and splenic CD4+ T cells expressed markedly increased levels of IL-4-specific message, but did not result in changes in IFN-gamma mRNA expression. To determine whether the route of immunization influenced IgE responses, mice were immunized s.c. with TT and CT as adjuvant. Significant increases in total and TT-specific IgE Abs were induced when CT was co-administered. Taken together, these results show that CT acts as a mucosal adjuvant to enhance Th2-type responses and in particular, the IL-4 produced results in a characteristic Ab isotype pattern associated with this cytokine.
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PMID:Mucosal adjuvant effect of cholera toxin in mice results from induction of T helper 2 (Th2) cells and IL-4. 759 61

A reverse transcriptase-PCR strategy was developed for the detection of hog cholera virus. Hog cholera virus template was amplified from tissue culture fluids and from tissues and blood of infected pigs, but not from samples containing other pestiviruses. Restriction endonuclease analysis identified samples as historic or recent isolates.
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PMID:Reverse transcriptase-PCR assay for detection of hog cholera virus. 781 9

Seven days after activation with concanavalin A and irradiated spleen cells, murine CD4+ T cells were re-stimulated with ionomycin and phorbol 12-myristate 13-acetate (PMA). IL-2 and IL-4 were determined in the supernatant. When cholera toxin, forskolin together with phosphodiesterase inhibitors or dibutyryl-cAMP were added at the time of re-stimulation, a dose-dependent increase of IL-4 and IL-5 release was noted. IL-2 was down-regulated as reported before. The up-regulation of IL-4 and the down-regulation of IL-2 correlated with an increase of IL-4 mRNA and a decrease of IL-2 mRNA as determined by semi-quantitative reverse transcriptase polymerase chain reaction. Similar results were found with prostaglandin E2 using PMA and ionomycin or plate-bound anti-CD3 antibody as re-stimulants. These results suggest that, in activated CD4+ T cells, cAMP-elevating agents induce a switch of lymphokine production towards a Th2-like phenotype through regulation at the transcriptional level. This is supported by the fact that complex formation between a synthetic nuclear factor of activated T cells (NF-AT) binding site from the IL-2 promoter and nuclear extracts was decreased when cholera toxin was added to re-activated CD4+ T cells, suggesting that cholera toxin and cAMP down-regulate IL-2 expression via decreased NF-AT binding. Finally, since IL-4 has been reported to amplify IL-4 release from activated CD4+ T cells, the autoinduction of IL-4 may very well function via cAMP.
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PMID:cAMP up-regulates IL-4 and IL-5 production from activated CD4+ T cells while decreasing IL-2 release and NF-AT induction. 781 41

We have used the potent mucosal immunogen cholera toxin (CT) to assess antigen-specific CD4+ T-cell responses, including Th1- and Th2-type cells in mucosa-associated tissues, e.g. Peyer's patches (PP), and systemic tissue, e.g. spleen (SP), for their regulatory role in the induction of CT-specific B-cell antibody responses in the gastrointestinal (GI) tract as well as in systemic sites. The CT was given by either oral or intravenous (i.v.) routes and the mice orally immunized with CT exhibited brisk IgA anti-CT antibody responses in faecal extracts and elevated IgG anti-CT antibody responses in serum. Further, significant IgA anti-CT spot-forming cells (SFCs) were seen in lamina propria lymphocytes (LPLs) from mice orally immunized with CT. In contrast, i.v. immunization with CT induced IgM and IgG anti-CT SFC responses in SP, and serum anti-CT antibodies of these two isotypes; no anti-CT responses were induced in the GI tract after immunization by this route. The CD4+ T cells isolated from PP and SP of mice orally immunized with CT were stimulated in vitro with CT-B-coated latex microspheres for 1-6 days, and the induction of IL-2 and interferon gamma (IFN-gamma) (Th1-type) or IL-4 and IL-5 (Th2-type) producing SFCs were analysed by a cytokine-specific ELISPOT and cytokine-specific mRNA was detected by reverse transcriptase (RT)-PCR assays.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Helper Th1 and Th2 cell responses following mucosal or systemic immunization with cholera toxin. 797 32

Although thyrotropin is known to regulate thyroid cell differentiation and proliferation, human thyroid carcinoma cells are relatively insensitive or resistant to TSH stimulation. The expression levels of TSH receptor are significantly lower in carcinoma tissues than in normal tissues. Furthermore, in vitro human thyroid cell growth is not regulated by TSH itself. We, therefore, isolated neomycin-resistant stable human thyroid carcinoma cell (WRO cell) transfectants overexpressing intact human TSH receptor to evaluate the functional role of TSH receptor on carcinoma cells. Southern blot analysis confirmed incorporation and amplification of human TSH receptor complementary DNA sequences into genomic DNA. Northern gel analysis and reverse transcriptase-polymerase chain reaction analysis revealed the presence of specific TSH receptor messenger RNA (4.0 kilobases), and the specific binding and the affinity of [125I]TSH on stably transfected WRO cells were demonstrated compared to wild type. Nevertheless, impaired cAMP production to transfectants by TSH was observed. cAMP production was confirmed after stimulation of both wild type and transfectants by forskolin, cholera toxin, and isoproterenol. In contrast, TSH could affect the cytoplasmic calcium mobilization immediately after the addition of TSH to WRO transfectants. These results suggest that the impairment of TSH action on human thyroid carcinoma cells is not due to a major structural abnormality of the TSH receptor, reduction in the receptor number, or receptor affinity, but much more likely due to a TSH receptor-guanyl nucleotide-binding protein coupling defect.
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PMID:Overexpression of the intact thyrotropin receptor in a human thyroid carcinoma cell line. 838 Oct 75

The phylogenetic relationships of 50 reference strains, mostly marine bacteria which require Na+ for growth, were determined on the basis of 600 16S rRNA nucleotides by using reverse transcriptase sequencing. Strains belonging to 10 genera were included (four genera of the family Vibrionaceae, the genus Aeromonas of the family Aeromonadaceae, and the genera Alteromonas, Marinomonas, Shewanella, Pseudomonas, and Deleya). The sequences were aligned, the similarity values and evolutionary distance values were determined, and a phylogenetic tree was constructed by using the neighbor-joining method. On the basis of our results, the family Vibrionaceae was separated into at least seven groups (genera and families). Vibrio marinus clearly was on a line of descent that was remote from other vibrios. As determined by the similarity and evolutionary distance values, V. marinus is more distantly related to the family Vibrionaceae than the members of the Aeromonadaceae are. Also, Vibrio cholerae strains formed a separate group with Vibrio mimicus at the genus level. Of 30 species of the Vibrionaceae, 17 formed a large phylogenetic cluster. The genus Listonella was found to be a heterogeneous group, and the species were distributed in various subgroups of the Vibrionaceae. The separation of the family Aeromonadaceae from the family Vibrionaceae and the separation of the genera Marinomonas and Shewanella from the genus Alteromonas were confirmed in this phylogenetic study. However, a marine Pseudomonas species, Pseudomonas nautica, was clearly separated from two terrestrial Pseudomonas species. Each group that was separated by the phylogenetic analysis had characteristic 16S rRNA sequence patterns that were common only to species in that group. Therefore, the characteristic sequences described in this paper may be useful for identification purposes.
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PMID:Phylogenetic relationships of marine bacteria, mainly members of the family Vibrionaceae, determined on the basis of 16S rRNA sequences. 842 11


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