Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.1 (RNase)
 16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The solution structure of Escherichia coli tRNA(3Thr) (anticodon GGU) and the residues of this tRNA in contact with the alpha 2 dimeric threonyl-tRNA synthetase were studied by chemical and enzymatic footprinting experiments. Alkylation of phosphodiester bonds by ethylnitrosourea and of N-7 positions in guanosines and N-3 positions in cytidines by dimethyl sulphate as well as carbethoxylation of N-7 positions in adenosines by diethyl pyrocarbonate were conducted on different conformers of tRNA(3Thr). The enzymatic structural probes were nuclease S1 and the cobra venom ribonuclease. Results will be compared to those of three other tRNAs, tRNA(Asp), tRNA(Phe) and tRNA(Trp), already mapped with these probes. The reactivity of phosphates towards ethylnitrosourea of the unfolded tRNA was compared to that of the native molecule. The alkylation pattern of tRNA(3Thr) shows some similarities to that of yeast tRNA(Phe) and mammalian tRNA(Trp), especially in the D-arm (positions 19 and 24) and with tRNA(Trp), at position 50, the junction between the variable region and the T-stem. In the T-loop, tRNA(3Thr), similarly to the three other tRNAs, shows protections against alkylation at phosphates 59 and 60. However, tRNA(3Thr) is unique as far as very strong protections are also found for phosphates 55 to 58 in the T-loop. Compared with yeast tRNA(Asp), the main differences in reactivity concern phosphates 19, 24 and 50. Mapping of bases with dimethyl sulphate and diethyl pyrocarbonate reveal conformational similarities with yeast tRNA(Phe). A striking conformational feature of tRNA(3Thr) is found in the 3'-side of its anticodon stem, where G40, surrounded by two G residues, is alkylated under native conditions, in contrast to other G residues in stem regions of tRNAs which are unreactive when sandwiched between two purines. This data is indicative of a perturbed helical conformation in the anticodon stem at the level of the 30-40 base pairs. Footprinting experiments, with chemical and enzymatic probes, on the tRNA complexed with its cognate threonyl-tRNA synthetase indicate significant protections in the anticodon stem and loop region, in the extra-loop, and in the amino acid accepting region. The involvement of the anticodon of tRNA(3Thr) in the recognition process with threonyl-tRNA synthetase was demonstrated by nuclease S1 mapping and by the protection of G34 and G35 against alkylation by dimethyl sulphate. These data are discussed in the light of the tRNA/synthetase recognition problem and of the structural and functional properties of the tRNA-like structure present in the operator region of the thrS mRNA.
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PMID:Tertiary structure of Escherichia coli tRNA(3Thr) in solution and interaction of this tRNA with the cognate threonyl-tRNA synthetase. 245

A combination of several enzymes, RNase-T1, nuclease S1, T4-polynucleotide kinase and T4-RNA ligase were used to prepare and modify different fragments of yeast tRNAAsp (normal anticodon G U C). This allowed us to reconstitute, in vitro, a chimeric tRNA that has any of the four bases G, A, U or C, as the first anticodon nucleotide, labelled with (32p) in its 3' position. Such reconstituted (32p) labelled yeast tRNAAsp were microinjected into the cytoplasm or the nucleus of the frog oocyte and checked for their stability as well as for their potential to work as a substrate for the maturation (modifying) enzymes under in vivo conditions. Our results indicate that the chimeric yeast tRNAsAsp were quite stable inside the frog oocyte. Also, the G34 was effectively transformed inside the cytoplasm of frog oocyte into Q34 and mannosyl-Q34; U34 into mcm5s2U and mcm5U. In contrast, C34 and A34 were not transformed at all neither in the cytoplasm nor in the nucleus of the frog oocyte. The above procedure constitutes a new approach in order to detect the presence of a given modifying enzyme inside the frog oocyte; also it provides informations about its cellular location and possibility about its specificity of interaction with foreign tRNA.
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PMID:Enzymatic replacement in vitro of the first anticodon base of yeast tRNAAsp: application to the study of tRNA maturation in vivo, after microinjection into frog oocytes. 628 19

To investigate further the presence of an autocrine proliferative loop involving gastrin in colorectal carcinomas and to clarify the receptor responsible, 102 human colorectal carcinomas and 10 hepatic metastases were investigated for the expression of the genes encoding gastrin, the gastrin/CCK-B receptor and the gastrin/CCK-C receptor. Levels of RNA expression were assayed by RNase protection assay. In addition, gastrin/CCK receptors on crude membranes of tumour tissue were assayed by radioligand binding. High-affinity gastrin/CCK-B receptors were not detected in any of the carcinomas investigated, whereas in 36% low-affinity binding was observed, consistent with the expression of the gastrin/CCK-C receptor. RNase protection assay detected the RNA for the gastrin/CCK-B receptor in 11% of the carcinomas investigated, whereas the RNA for the gastrin/CCK-C receptor was demonstrated in 75% and the RNA for gastrin in 86% of the carcinomas investigated. These results confirm the recent demonstration of progastrin fragments in colorectal carcinomas. One possible explanation for progastrin expression is that such progastrin fragments may participate in an autocrine proliferative loop. The receptor involved in this loop is more likely to be the low-affinity gastrin/CCK-C receptor rather than the gastrin/CCK-B receptor, which is rarely expressed in colorectal carcinomas.
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PMID:Expression of gastrin, gastrin/CCK-B and gastrin/CCK-C receptors in human colorectal carcinomas. 759 30

Gastrin is transcriptionally responsive to EGF stimulation (Merchant et al., 1991, Mol. Cell. Biol., 11:2686-2696). Consequently, we hypothesized that previously recognized gastrin autocrine loops (Hoosein et al., 1990, Exp. Cell. Res., 186:15-21), might be controlled by autocrine TGF alpha in human colon carcinoma cells. Therefore, we examined the interaction between these two autocrine growth factors in two colon carcinoma cell lines which utilize TGF alpha. The FET cell line requires exogenous TGF alpha/EGF for optimal growth and has a classical TGF alpha autocrine loop which is disrupted by TGF alpha or epidermal growth factor receptor (EGFr) antibodies. The HCT 116 cell line is not dependent on exogenous TGF alpha/EGF and exhibits a nonclassical TGF alpha autocrine loop which is not disrupted by neutralizing antibodies to either TGF alpha itself or the EGFr. Basal gastrin mRNA production is significantly higher in HCT 116 than FET as measured by RNase protection assay. In the FET cells, exogenous EGF stimulates gastrin mRNA production but not in HCT 116. When the TGF alpha autocrine loop in HCT 116 is disrupted by constitutive expression of antisense TGF alpha mRNA, the gastrin mRNA level is significantly repressed. In xenografts derived from these antisense clones, TGF alpha reverted to high expression, and the gastrin mRNA level was again increased. This interaction between the strong TGF alpha loop in HCT 116 and the gastrin autocrine loop may confer a growth advantage to these colon cells. Such interactions between growth factors may promote enhanced tumorigenicity to transformed cells with these strong, nonclassical autocrine loops.
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PMID:Regulation of autocrine gastrin expression by the TGF alpha autocrine loop. 782 34

Gastrin, produced in the G-cells of the gastric antrum and regulating acid secretion in the stomach, also acts as a trophic factor in the gastrointestinal tract. Because of its possible role in colon cell proliferation and differentiation, evidence for its presence in normal colorectal mucosa and adenocarcinoma was sought. Utilizing tumors and matched normal mucosa from 26 patients, mature gastrin and progastrin were studied by immunohistochemistry. In normal colonic mucosal crypts, occasional cells stained concordantly for gastrin, progastrin, and chromogranin A, suggesting that they are of neuroendocrine origin. Adenomatous polyps stained neither for gastrin nor chromogranin A. In 22 of 23 adenocarcinomas, more than 50% of tumor cells stained for gastrin and progastrin. The expected gastrin transcript was demonstrable by polymerase chain reaction and RNase protection in tumors and by polymerase chain reaction in normal mucosa. Its identity was confirmed by sequencing the polymerase chain reaction product. A larger transcript containing Intron II was present in both cancers and normal mucosa but was barely discernible in the gastric antrum. Aberrant expression of gastrin may contribute to deregulated proliferation of many colorectal carcinomas.
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PMID:Expression of the gastrin gene in the normal human colon and colorectal adenocarcinoma. 850 33

The present study was designed to investigate whether heparin-binding epidermal growth factor-like growth factor and its related peptides are expressed in response to gastrin in rat stomach. Rat gastrin-17I (2.5 nmol/kg/hour) or gastrin-17I plus gastrin receptor antagonist, L-740,093 (2.0 mg/kg/hour), was injected intravenously into male Sprague-Dawley rats. RNA was extracted from oxyntic mucosa, and heparin-binding epidermal growth factor-like growth factor and related peptide gene expression was estimated using a ribonuclease protection assay. The level of transforming growth factor-alpha mRNA did not change at any time point during the experiment. In contrast, the levels of heparin-binding epidermal growth factor-like growth factor and amphiregulin mRNA were significantly increased within 3 hours following gastrin infusion and reached maximum levels 6 and 12 hours later, respectively. Continuous infusion of gastrin significantly increased oxyntic mucosal proliferation. Gastrin receptor antagonist significantly inhibited gastrin-induced heparin-binding epidermal growth factor-like growth factor and amphiregulin gene expression and gastrin-induced oxyntic mucosal proliferation. These findings indicate that heparin-binding epidermal growth factor-like growth factor and amphiregulin genes are induced by gastrin and that they play a role in the trophic action of gastrin on oxyntic mucosa.
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PMID:Induction of heparin binding epidermal growth factor-like growth factor and amphiregulin mRNAs by gastrin in the rat stomach. 920 88

1. Inhibition of gastric acid secretion by proton pump inhibitors like omeprazole increases the synthesis and secretion of the pyloric antral hormone gastrin. We report here how omeprazole influences the conversion of the gastrin precursor to its final products, and the abundance of mRNAs encoding proteins associated with progastrin processing in rat antral mucosa. 2. Progastrin processing was studied using a pulse-chase protocol in antral mucosa, incubated in vitro, from rats treated with omeprazole for up to 5 days. Labelled peptides were detected by on-line scintillation counting after immunoprecipitation and HPLC. The mRNAs encoding prohormone-processing enzymes were identified by Northern blot, polymerase chain reaction or ribonuclease protection assay, and their cellular origins identified by immunocytochemistry. 3. Cleavage of [3H]- and [35S]-labelled progastrins at Arg-94-95 or Arg-57-58, and amidation at Phe-92 were not influenced by pretreatment with omeprazole. In contrast, cleavage of G34 (the thirty-four amino acid amidated gastrin) at Lys-74-75 to give G17 (the seventeen amino acid amidated gastrin), and of G34-Gly to G17-Gly (G34 and G17 with COOH-terminal glycine), was increased 3-fold after treatment with omeprazole for either 1 or 5 days. 4. Approximately 20% of newly synthesized amidated and Gly-extended gastrins were secreted within 240 min of the labelling period in omeprazole-treated samples, but secretion of labelled gastrins from control tissue was undetectable over a comparable period. 5. The amidating enzyme, peptidyglycine alpha-amidating mono-oxygenase (PAM), the prohormone convertases PC1/3, PC2, PC5 and the PC2 chaperone 7B2 were localized to rat antral gastrin cells by immunocytochemistry. The relative abundance of mRNA species encoding 7B2, PC5 and PAM were unchanged after treatment with omeprazole for 5 days, whereas gastrin, PC1/3 and PC2 mRNAs are known to increase at this time. 6. The main consequence of increased cleavage at Lys-74-75 is the production of G17 and G17-Gly at the expense of G34 and G34-Gly, respectively. The latter have longer plasma half-lives, and so their increased cleavage may serve to limit the rise in plasma gastrin concentration after inhibition of acid secretion. Changes in the abundance of mRNAs encoding prohormone-processing enzymes cannot account for the rapidity of the changes in cleavage of progastrin at Lys residues after omeprazole.
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PMID:Regulation by gastric acid of the processing of progastrin-derived peptides in rat antral mucosa. 926 20

Recent studies have suggested that cholecystokinin (CCK) receptors may play a role in the development and growth of pancreatic cancers. We detected the expression of mRNA encoding CCK-A and CCK-B receptors in eight human pancreatic tumour cell lines using reverse transcription-polymerase chain reaction (RT-PCR), but not by RNase protection assays. The K-ras gene, which can be activated by G-coupled protein receptors such as CCK receptors, was mutated in codon 12 in five of the cell lines. In addition, Mia PaCa-2 pancreatic cancer cells did not respond to CCK or gastrin in cell proliferation or focal adhesion kinase (FAK) phosphorylation assays. In contrast, mouse NIH3T3 fibroblasts transfected with human CCK-B receptor (NIH3T3CCK-BR) showed increased proliferation and phosphorylation to the peptides. Also, radioligand binding studies indicated that Mia PaCa-2 cells had approximately 12.5-fold less CCK-B receptors than NIH3T3CCK-BR. Our results suggest that in Mia PaCa-2 cells, CCK receptors may not play a crucial role in supporting cell growth.
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PMID:Cholecystokinin receptors in human pancreatic cancer cell lines. 984 31

The murine anti-bombesin monoclonal antibody, 2A11, has been demonstrated to inhibit growth of some small-cell lung cancer (SCLC) cells in nude mice xenografts and in a clinical trial. To determine if the expression of bombesin-like peptides (BLP) and their receptors (GRP-R and NMB-R) correlate with an in vitro response to 2A11, we measured these parameters in seven SCLC cell lines. Gastrin releasing peptide (GRP) mRNA was detected in three of seven cell lines (NCI-H69, NCI-H345, NCI-H510) and neuromedin B (NMB) mRNA was detected in all seven lines using an RNase protection assay (RPA). Immunoreactive BLP was detected in the cell pellets of all lines (range 0.11-59.90 pmol/mg protein) by a solid phase GRP radioimmunoassay (RIA) using 125I-labeled 2A11. RPA detected GRP-receptor mRNA in two cell lines (NCI-H69 and NCI-H345) and NMB-receptor in three lines (NCI-H345, NCI-H510, and NCI-H660). Reverse transcriptase-PCR confirmed the presence of receptor mRNA in these lines and detected NMB-receptor in an additional three lines (NCI-H69, NCI-H82, and NCI-H187). Calcium mobilization in response to BLP stimulation was detected in the six cell lines expressing either GRP-R or NMB-R mRNA but not in NCI-N417, which had no detectable BLP-receptor. 2A11 (5 microg/ml) inhibited colony formation by 26-61% after 2 weeks in all cell lines except NCI-N417. Thus, growth inhibition by 2A11 requires the presence of at least one BLP-receptor. These findings may be useful in selecting patients with SCLC for treatment with 2A11.
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PMID:Correlation of expression of bombesin-like peptides and receptors with growth inhibition by an anti-bombesin antibody in small-cell lung cancer cell lines. 985 94

Chronic inflammation of the gastric epithelium is believed to induce mucosal changes that can eventually develop into gastric cancer. In gastrin-deficient (G-/-) mice exhibiting chronic inflammation in the hypochlorhydric stomach, we documented a prominent fundic mucous cell lineage sharing morphological similarity with preneoplastic changes reported in Helicobacter-infected mice. To study the identity and origin of this cell lineage, we screened for different gastric mucosal cell markers. The clusters of large, foamy cells stained for trefoil factor 2 (TFF2/SP), MUC6 and the lectin Griffonia Simplicifolia II (GSII), but not for the intestine-specific transcription factor Cdx2, suggested that they arise from gastric mucous neck cells. Ki67-labeled GSII-positive neck cells in Helicobacter felis-infected, but not G-/- stomachs, suggested that mucous neck cell proliferation accounted for expansion of this compartment in the H. felis model of gastritis, but not the G-/- model. Using RNase protection assays and quantitative PCR, we found that interferon gamma (IFNgamma) was the most abundant proinflammatory cytokine in the G-/- stomach. We also found that this Th1 cytokine can increase the abundance of mucous neck cells, since its infusion into mice recapitulated the appearance of these cells as observed in both G-/- and H. felis-infected mice. Using the human gastric cell line NCI-N87, we showed that IFNgamma induces the secretion of mucus and expression of MUC6, TFF2 and pepsinogen II, but not of pepsinogen I and intrinsic factor. In conclusion, our results demonstrate that inflammation, specifically the proinflammatory cytokine IFNgamma, induced expansion of the fundic mucous neck cell compartment, which likely represents both increased mucus production and cell number.
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PMID:Interferon gamma induction of gastric mucous neck cell hypertrophy. 1576 19


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