UNIPROT:P01350 - FACTA Search

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Query: UNIPROT:P01350 (gastrin)
9,683 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We report below on the NMR structural characterization of the complex between AMP and a 40-mer RNA aptamer in aqueous solution. Resonance assignments are based on multinuclear multidimensional NMR studies on complexes uniformly 13C, 15N-labeled with either AMP or the RNA aptamer. AMP binds to an internal loop (labeled G7-G8-A9-A10-G11-A12-A13-A14-C15-U16-G17) and bulge (G34 positioned opposite the internal loop) segment in the RNA aptamer, and our NMR study provides insights into features of the RNA folding topology and the molecular recognition events in the AMP binding pocket on the RNA. Specifically, the helical stems are extended by G-G mismatch formation from either direction into the internal loop/bulge segment of the RNA aptamer on complex formation. The internal loop adopts a unique fold with the purine ring of AMP intercalated between A10 and G11 in the complex. The G8-A9-A10-AMP segment adopts certain stacking features in common with a GNRA turn and is closed by the G7.G11 mismatch pair. The purine rings of A12 and G34 (syn) are stacked on each other and participate in stablizing the AMP intercalation site. A large number of intermolecular NOEs have been identified between the AMP ligand and the G8, A10, G11, G17, U18, and G34 residues on the RNA aptamer in the complex. The Watson-Crick edge of the AMP is oriented toward the exocyclic amino group of G8, suggestive of a hydrogen-bonding alignment between G8 and AMP in the complex. The AMP sugar ring is positioned in the minor groove of the rightward helical stem centered about the G17.G34 mismatch and U18.A33 Watson-Crick pairs. The AMP binds to one face of the folded internal loop/bulge segment of the RNA aptamer while the opposite face is capped by a stacked alignment of the A13-A14-C15-U16 segment located toward the 3'-end of the internal loop segment. Globally, the two helical stems of the RNA aptamer are aligned approximately orthogonal to each other with tertiary interactions centered about the internal loop/bulge segment generating the AMP binding site on the RNA.
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PMID:RNA folding topology and intermolecular contacts in the AMP-RNA aptamer complex. 885 64

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

Gastric endocrine cell populations and serum and tissue gastrin have been examined in sheep which were infected either intraruminally by tube with 150,000 Ostertagia circumcincta larvae followed by a trickle infection of 10,000 larvae thrice weekly for 8 weeks or by the transfer of 15,000 adult worms directly into the abomasum and killed 8 days later. Depletion of both antral gastrin and somatostatin was evident in both groups: tissue gastrin concentrations were reduced by 85% in the trickle infection and both G cells (gastrin-containing) and D cells (somatostatin-containing) were pale and fewer after adult worm transfer. The concurrent depletion of antral gastrin and somatostatin supports the contention that the hypergastrinaemia in parasitised sheep is largely secondary to the increase in abomasal pH. Although there was no change in the proportions of G34 and G17 in the tissues, there was an increase in the longer form of gastrin in the circulation of the larval-infected sheep, suggesting that there may be differential secretion of G17 and G34 which may be exaggerated as the rate of secretion increases. Although the fundic mucosa was thicker following trickle infection, there was no evidence of enterochromaffin-like cell hyperplasia in either infected group. It is suggested that hyper-gastrinaemia may be beneficial to the host, as it may allow the abomasum to regain the ability to acidify its contents during continued exposure to the parasites.
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PMID:Infection of sheep with adult and larval Ostertagia circumcincta: gastrin. 977 Jun 25

We previously observed that the trophic actions of gastrin (G17) on the AR42J rat acinar cell line are mediated by mitogen-activated protein kinase (MAPK)-induced c-fos gene transcription via protein kinase C (PKC)-dependent and -independent pathways. In this study, we further investigated the signaling pathways that target c-fos in response to G17. G17 led to a sixfold induction in luciferase activity in cells transfected with plasmids containing the -356+109 sequence of the murine c-fos promoter, which includes the Sis-inducible element (SIE), serum response element (SRE), and the Ca2+/cAMP response element (CRE) regulatory elements. Addition of either the selective PKC inhibitor GF-109203X or the MAPK/extracellular signal-regulated kinase inhibitor PD-98059 resulted in an 80% reduction in luciferase activity. G17 induced the transcriptional activity of both Elk-1 and Sap-1a, transcription factors that bind to the E26 transformation specific (Ets) DNA sequence of the SRE, and this effect was inhibited by both GF-109203X and PD-98059. Point mutations in the Ets sequence led to a 4-fold induction of c-fos transcription stimulated by G17 and to a 1.3-fold induction in response to epidermal growth factor (EGF). In contrast, mutations in the CA rich G (CArG) sequence of the SRE prevented transcriptional activation by both G17 and EGF. G17 induction of the Ets mutant construct was unaffected by either GF-109203X or PD-98059. Because activation of the SRE involves the small GTP-binding protein Rho A, we examined the role of Rho A in G17 induction of c-fos transcription. Inactivation of Rho A by either the specific inhibitor C3 or by expression of a dominant negative Rho A gene inhibited G17 induction of both the wild-type and the Ets mutant constructs by 60%. C3 also inhibited G17-stimulated AR42J cell proliferation. Thus G17 targets the c-fos promoter CArG sequence via Rho A-dependent pathways, and Rho A appears to play an important role in the regulation of the trophic action of G17.
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PMID:Gastrin induces c-fos gene transcription via multiple signaling pathways. 995 Aug 15

The therapeutic effect of antibodies raised by the immunogen Gastrimmune was compared with both a CCKB/gastrin receptor antagonist, CI-988, and 5-Fluorouracil/leucovorin in a gastric cancer model. The human gastric ascites cell line, MGLVA1asc, produced and secreted progastrin and glycine-extended gastrin as determined by radioimmunoassay and immunocytochemistry. Cells were also stained with an antiserum directed against the human CCKB/gastrin receptor. MGLVAI asc cells were injected i.p. into SCID mice. Antibodies raised by Gastrimmune immunization of rabbits (affinity for G17 of 0.15 nM and GlyG17 of 0.47 nM) were passively infused i.p. and significantly enhanced survival by up to 5 days (p=0.0024 from vehicle controls). The enhancement in survival was not significantly different from that achieved by treatment with 5-Fluorouracil and leucovorin. A CCKB/gastrin receptor antagonist, CI-988, did not affect survival with cells injected at 7.5 x 10(5) cells/mouse but significantly increased the survival of mice injected with a lower cell innoculum of 5 x 10(5) cells/mouse from 30 to 35 days (p=0.0186). At this lower innoculum antibodies raised by Gastrimmune induced complete survival in 2 animals with the remaining dead by day 36 (p=0.0022). Thus, both endocrine and autocrine pathways mediated by precursor and mature gastrin molecules may be jointly operational in the gastric cancer scenario and may be important targets for therapeutic agents.
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PMID:A comparison of an anti-gastrin antibody and cytotoxic drugs in the therapy of human gastric ascites in SCID mice. 1018 27

1. Gastrointestinal endocrine cells produce biogenic amines which are transported into secretory vesicles by one of two proton-amine exchangers, vesicular monoamine transporters type 1 and 2 (VMAT1 and 2). We report here the presence of VMAT1 in rat gastrin (G) cells and the relevance of VMAT1 function for the modulation of progastrin processing by biogenic and dietary amines. 2. In immunocytochemical studies VMAT1, but not VMAT2, was localized to subpopulations of G cells and enterochromaffin (EC) cells; neither was found in antral D cells. The expression of VMAT1 in antral mucosa was confirmed by Northern blot analysis, which revealed an mRNA band of approximately 3.2 kb, and by Western blot analysis, which revealed a major protein of 55 kDa. 3. In pulse-chase labelling experiments, the conversion of the amidated gastrin G34 to G17 was inhibited by biogenic amine precursors (L-DOPA and 5-hydroxytryptophan). This inhibition was stereospecific and sensitive to reserpine (50 nM), which blocks VMAT1 and VMAT2, but resistant to tetrabenazine, which is a selective inhibitor of VMAT2. 4. Dietary amines such as tyramine and tryptamine also inhibited G34 cleavage. This effect was associated with a loss of the electron-dense core of G cell secretory vesicles. It was not stereospecific or reserpine sensitive, but was correlated with hydrophobicity. 5. Thus rat antral G cells can express VMAT1; transport of biogenic amines into secretory vesicles by VMAT1 is associated with inhibition of G34 cleavage, perhaps by raising intravesicular pH. Dietary amines also modulate cleavage of progastrin-derived peptides, but do so by a VMAT1-independent mechanism; they may act as weak bases that passively permeate secretory vesicle membranes and raise intravesicular pH.
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PMID:Modulation of gastrin processing by vesicular monoamine transporter type 1 (VMAT1) in rat gastrin cells. 1033 97

Gastrin (G17) has a CCKB receptor-mediated growth-promoting effect on the AR42J rat acinar cell line that is linked to induction of both mitogen-activated protein kinase (MAPK) and c-fos gene expression. We investigated the mechanisms that regulate the growth factor action of G17 on the rat pituitary adenoma cell line GH3. Both AR42J and GH3 cells displayed equal levels of CCKB receptor expression and similar binding kinetics of 125I-labeled G17. G17 stimulation of cell proliferation was identical in both cell lines. G17 stimulation of GH3 cell proliferation was completely blocked by the CCKB receptor antagonist D2 but not by the MEK inhibitor PD-98059 or the protein kinase C inhibitor GF-109203X, which completely inhibited G17 induction of AR42J cell proliferation. G17 induced a c-fos SRE-luciferase reporter gene plasmid more than fourfold in the AR42J cells, whereas it had no effect in the GH3 cells. In contrast to what we observed in the AR42J cells, G17 failed to stimulate MAPK activation and Shc tyrosyl phosphorylation and association with the adapter protein Grb2. Epidermal growth factor induced the MAPK pathway in the GH3 cells, demonstrating the integrity of this signaling system. G17 induced Ca2+ mobilization in both the GH3 and AR42J cells. The calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide inhibited AR42J cell proliferation by 20%, whereas it completely blocked G17 induction of GH3 cell growth. The Ca2+ ionophore ionomycin stimulated GH3 cell proliferation to a level similar to that observed in response to G17, but it had no effect on AR42J cell proliferation. Thus there are cell type specific differences in the requirement of the MAPK pathway for the growth factor action of G17. Whereas in the AR42J cells G17 stimulates cell growth through activation of MAPK and c-fos gene expression, in the GH3 cells, G17 fails to activate MAPK, and it induces cell proliferation through Ca2+-dependent signaling pathways. Furthermore, induction of Ca2+ mobilization in the AR42J cells appears not to be sufficient to sustain cell proliferation.
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PMID:Cell type-specific requirement of the MAPK pathway for the growth factor action of gastrin. 1036 39

Posttranslational processing of progastrin to a carboxy terminally amidated form (G-NH(2)) is essential for its effect on gastric acid secretion and other biological effects mediated by gastrin/CCK-B receptors. The immediate biosynthetic precursor of G-NH(2), glycine-extended gastrin (G-Gly), does not stimulate gastric acid secretion at physiological concentrations but is found in high concentrations during development. G-NH(2) and G-Gly have potent growth stimulatory effects on gastrointestinal tissues, and G-NH(2) can stimulate proliferation of human kidney cells. Thus we sought to explore the actions of G-NH(2) and G-Gly on the human embryonic kidney cell line HEK 293. HEK 293 cells showed specific binding sites for (125)I-labeled Leu(15)-G17-NH(2) and (125)I-Leu(15)-G(2-17)-Gly. Both G-NH(2) and G-Gly induced a dose-dependent increase in [(3)H]thymidine incorporation, and both peptides together significantly increased [(3)H]thymidine incorporation above the level of either peptide alone. G-NH(2) and G-Gly were detected by radioimmunoassay in serum-free conditioned media. Antibodies directed against G-NH(2) and G-Gly lead to a significant reduction in [(3)H]thymidine incorporation. G-NH(2) but not G-Gly increased intracellular Ca(2+) concentration. We conclude that G-NH(2) and G-Gly act cooperatively via distinct receptors to stimulate the growth of a nongastrointestinal cell line (HEK 293) in an autocrine fashion.
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PMID:Glycine-extended gastrin regulates HEK cell growth. 1044 66

Gastrin is initially synthesized as a large precursor that requires endoproteolytic cleavage by a prohormone convertase (PC) for bioactivation. Gastric antral G-cells process progastrin at Arg(94)Arg(95) and Lys(74)Lys(75) residues generating gastrin heptadecapeptide (G17-NH(2)). Conversely, duodenal G-cells process progastrin to gastrin tetratriacontapeptide (G34-NH(2)) with little processing at Lys(74)Lys(75). Both tissues express PC1/PC3 and PC2. Previously, we demonstrated that heterologous expression of progastrin in an endocrine cell line that expresses PC1/PC3 and little PC2 (AtT-20) resulted in the formation of G34-NH(2). To confirm that PC1/PC3 was responsible for progastrin processing in AtT-20 cells and capable of processing progastrin in vivo we coexpressed either human wild-type (Lys(74)Lys(75)) or mutant (Arg(74)Arg(75), Lys(74)Arg(75), and Arg(74)Lys(75)) progastrins in AtT-20 cells with two different antisense PC1/PC3 constructs. Coexpression of either antisense construct resulted in a consistent decrease in G34-NH(2) formation. Gastrin mRNA expression and progastrin synthesis were equivalent in each cell line. Although mutation of the Lys(74)Lys(75) site within G34-NH(2) to Lys(74)Arg(75) resulted in the production of primarily G17-NH(2) rather than G34-NH(2), inhibition of PC1/PC3 did not significantly inhibit processing at the Lys(74)Arg(75) site. We conclude that PC1/PC3 is a progastrin processing enzyme, suggesting a role for PC1/PC3 progastrin processing in G-cells.
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PMID:Diminished prohormone convertase 3 expression (PC1/PC3) inhibits progastrin post-translational processing. 1077 9

Epidermal growth factor (EGF) and transforming growth factor-alpha (TGF-alpha) increase transcription of the gastrin gene, and the gastrin peptide may be phosphorylated by EGF-stimulated tyrosine kinase. Our aims were to compare EGF/TGF-alpha interactions in 2 human gastro-intestinal cell lines: MGLVA1, with a strong gastrin autocrine pathway, and C170HM2, with a weak pathway. Both cell lines expressed the TGF-alpha gene. MGLVA1 expressed TGF-alpha protein as determined by immuno-cytochemistry, which was absent in C170HM2. Both cell lines expressed the same level of EGF receptors, as assessed by flow cytometry; however, MGLVA1 did not have enhanced in vitro proliferation in response to EGF or TGF-alpha, unlike C170HM2. The basal growth of MGLVA1 was inhibited by anti-sera against TGF-alpha, the EGF receptor and G17. C170HM2 was not inhibited by any of the anti-sera. Neutralisation of TGF-alpha resulted in undetectable cell-associated progastrin levels in MGLVA1 (untreated had 391.7 fmol/5 x 10(6) cells). The progastrin level of C170HM2 remained unaffected. Tyrosine kinase activity, as assessed by phosphopeptide concentration, of unstimulated MGLVA1 was 2.6 times higher than that of C170HM2 in the cell membrane fraction (0.097 compared to 0.037 microg/mg protein, p < 0.001) and 4.8 times higher in the cytosolic fraction (0.269 compared to 0.056 microg/mg protein, p < 0.05). Following treatment with EGF, the phosphopeptide concentration increased in both the membrane and cytosolic fractions of both cell lines. Tyrphostin B42, which inhibits autophosphorylation of the EGF receptor, inhibited the basal growth of MGLVA1 (IC(50) 1.3 microM) and C170HM2 (9.5 microM, p < 0.05 from MGLVA1). Herbimycin, which inhibits pp60(c-src) kinase, reduced the basal growth of MGLVA1 (0.67 microM) but not C170HM2. Immunofluorescence studies confirmed the presence of tyrosine-phosphorylated proteins and pp60(c-src) within the cytoplasm of unstimulated MGLVA1 cells. There was no specific immunofluorescence for either parameter in C170HM2 cells until after treatment with EGF.
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PMID:Transforming growth factor-alpha-mediated growth pathways in human gastro-intestinal cell lines in relation to the gastrin autocrine pathway. 1086 48

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