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:4.6.1.1 (adenylate cyclase)
19,190 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have examined the conversion of parathyroid hormone-related peptide (PTHRP) from its NH2-terminally extended precursor pro-PTHRP. Within pro-PTHRP, an amino acid sequence exists that can serve as a substrate for the prohormone convertase, furin. To evaluate the potential role of furin in processing of this entity, we expressed pro-PTHRP in COS-7 cells, which normally produce this enzyme. Transiently transfected COS-7 cells secreted high levels of PTHRP into conditioned culture medium. Cotransfection of these cells with antisense furin cDNA resulted in marked inhibition of furin mRNA expression and secretion of an NH2-terminal fragment of pro-PTHRP, which comigrated with synthetic pro-PTHRP-(-12-->+36) on gel-permeation high-pressure liquid chromatography. In an adenylate cyclase bioassay, condition medium containing this fragment and synthetic pro-PTHRP-(-12-->+36) both exhibited lower potency than synthetic PTHRP-(1-36) and conditioned medium containing PTHRP produced by COS-7 cells in the absence of antisense furin. These results demonstrate the capacity of furin to convert pro-PTHRP to a more active product and suggest a role for this enzyme in the normal intracellular processing of this hormone.
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PMID:Processing of pro-PTHRP by the prohormone convertase, furin: effect on biological activity. 753 17

To attain full biological activity, the precursor pro-parathyroid hormone-related peptide (ProPTHRP) must be converted to the mature peptide PTHRP. We have examined the effect of inhibiting expression of the pro-hormone convertase furin in H-500 rat Leydig tumor cells on PTHRP production and action in vitro and in vivo. H-500 Leydig tumor cells were stably transfected with a mammalian expression plasmid containing furin cDNA in an anti-sense orientation. This resulted in inhibition of endogenous furin mRNA expression and of protein production as assessed by immunocytochemistry. These experimental cells secreted extended NH2-terminal PTHRP forms with reduced adenylate cyclase-stimulating activity. This was associated with a marked decrease in the proliferation of these tumor cells in vitro. Transfected and control cells were then implanted into male Fischer rats. Animals implanted with control cells became hypercalcemic. In contrast, animals implanted with experimental cells maintained near normal levels of plasma calcium. Experimental cells inoculated in vivo developed into tumors of significantly decreased volume compared to control cells and animal survival time was prolonged. Our results indicate that alteration of the processing of PTHRP can diminish the hypercalcemic endocrine actions of PTHRP and can reduce autocrine/paracrine effects of PTHRP on tumor cell growth both in vitro and in vivo. Furin may also exert a broader role in processing other factors required for tumor proliferation. Consequently, anti-sense modulation of furin activity may be a potential modality for understanding the mechanism of neoplastic growth and progression.
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PMID:Inhibition of processing of parathyroid hormone-related peptide by anti-sense furin: effect in vitro and in vivo on rat Leydig (H-500) tumor cells. 759 Dec 17

Prepro-vasoactive intestinal peptide (prepro VIP) was expressed in NIH 3T3 cells, and the prepro VIP-derived peptides produced by the cells were analyzed by chromatography combined with sequence-specific radio-immunoanalysis. In accordance with what has previously been reported on processing in non-endocrine cell lines, the VIP precursor was processed poorly in these non-endocrine cells. Mainly an extended form of VIP could be detected in the media from the cells, and no immunoreactivity specific for amidated VIP was found. However, by changing the dibasic cleavage site positioned N-terminal to the VIP sequence in the precursor into the consensus sequence (Arg, X,Lys/Arg,Arg) for the ubiquitous processing enzyme furin, thought to process, e.g. insulin receptors, factor VII, and by deleting residues 156-170 in the VIP precursor, expression of amidated VIP was obtained in this fibroblast cell line. Peptides from the wild-type VIP precursor as well as peptides from the mutated VIP precursor were found to be able to stimulate the adenylate cyclase in cells expressing the VIP receptor.
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PMID:Expression and characterization of VIP and two VIP mutants in NIH 3T3 cells. 813 20

Previously, we demonstrated that a single histamine H2 receptor can couple to both the adenosine 3',5'-cyclic monophosphate and inositol 1,4,5-trisphosphate/intracellular Ca2+ signaling pathways in a stimulatory manner. We undertook the present studies to fur her characterize the postreceptor events involved in H2 receptor dual signaling. Histamine H2 receptor-mediated signal transduction was examined in isolated cell membranes prepared from purified canine parietal cells and HEPA cells (rat hepatoma cell line) stably transfected to express the canine H2 histamine receptor cDNA. Histamine dose-dependently stimulated both adenylate cyclase [AC; mean effective concentration (EC50) = 2 x 10(-7) M] and phospholipase C (PLC; EC50 = 3.1 +/- 0.5 x 10(-7) M) activity in an H2-specific and GTP-dependent manner. Cholera toxin pretreatment abolished the stimulatory effect of histamine on PLC activity in isolated membranes without altering binding of the H2 receptor antagonist tiotidine. Anti-Gs alpha dose-dependently inhibited histamine-stimulated AC activity while leaving the effect of this secretagogue on PLC activity unaltered. Although anti-Gq alpha inhibited vasopressin-stimulated PLC activity in HEPA cells and carbachol-stimulated PLC in parietal cells, this antibody did not alter the action of histamine on PLC in the same membrane preparations. Antibody against the NH2 and COOH terminals of the common beta-subunit of heterotrimeric G proteins did not inhibit histamine-stimulated PLC activity. Our studies demonstrate for the the first time that activation of the H2 receptor leads to stimulation of both AC and PLC via separate GTP-dependent mechanisms.
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PMID:Histamine H2 receptor activates adenylate cyclase and PLC via separate GTP-dependent pathways. 889 80

The ocular ciliary epithelium is a bilayer of neuroepithelial cells specialized in the secretion of aqueous humor fluid and the regulation of intraocular pressure. In this study, we report on the expression of the regulatory peptide neurotensin (NT) and a set of differentiated neuroendocrine markers including neurotensin receptors (NTrs), the prohormone convertases furin, PC1, and PC2, and the neuroendocrine polypeptide 7B2 in the ciliary epithelium. Using a human cell line, ODM-2, derived from the nonpigmented ciliary epithelium, we demonstrate that (1) NT expression is highly activated by nerve growth factor, glucocorticoid, and activators of adenylate cyclase; (2) NTr expression is up-regulated by selective ligand-activated beta2-adrenergic receptor; and (3) PC1 and PC2 expression are up-regulated via distinct signaling transduction pathways. PC1 gene expression is activated by phorbol ester, and PC2 by the same inducers as those of NT expression. A radioimmunoassay for NT detected an NT-like immunoreactivity in human ciliary epithelium and ODM-2 cell extracts, in aqueous humor, and in conditioned culture medium. The results support the view that the entire ciliary epithelium functions as a neuroendocrine tissue, synthesizing, processing, and releasing NT into the aqueous humor where it may exert important physiological functions through autocrine and/or paracrine mechanisms.
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PMID:Molecular characterization and differential gene induction of the neuroendocrine-specific genes neurotensin, neurotensin receptor, PC1, PC2, and 7B2 in the human ocular ciliary epithelium. 934 25

Anthrax is primarily a disease of herbivores caused by gram-positive, aerobic, spore-forming Bacillus anthracis. Humans are accidental hosts through the food of animal origin and animal products. Anthrax is prevelant in most parts of the globe, and cases of anthrax have been reported from almost every country. Three forms of the disease have been recognized: cutaneous (through skin), gastrointestinal (through alimentary tract), and pulmonary (by inhalation of spores). The major virulence factors of Bacillus anthracis are a poly-D glutamic acid capsule and a three-component protein exotoxin. The genes coding for the toxin and the enzymes responsible for capsule production are carried on plasmid pXO1 and pXO2, respectively. The three proteins of the exotoxin are protective antigen (PA, 83 kDa), lethal factor (LF, 90 kDa), and edema factor (EF, 89 kDa). The toxins follow the A-B model with PA being the B moeity and LF/EF, the alternative A moeities. LF and EF are individually nontoxic, but in combination with PA form two toxins causing different pathogenic responses in animals and cultured cells. PA + LF forms the lethal toxin and PA + EF forms the edema toxin. During the process of intoxication, PA binds to the cell surface receptor and is cleaved at the sequence RKKR (167) by cell surface proteases such as furin generating a cell-bound, C-terminal 63 kDa protein (PA63). PA63 possesses a binding site to which LF or EF bind with high affinity. The complex is then internalized by receptor-mediated endocytosis. Acidification of the vesicle leads to instertion of PA63 into the endosomal membrane and translocation of LF/EF across the bilayer into the cytosol where they exert their toxic effects. EF has a calcium- and calmodulin-dependent adenylate cyclase activity. Recent reports indicate that LF is a protease that cleaves the amino terminus of mitogen-activated protein kinase kinases 1 and 2 (MAPKK1 and 2), and this cleavage inactivates MAPKK1 and thus inhibits the mitogen-activated protein kinase signal transduction pathway. We describe in detail the studies so far done on unraveling the molecular mechanisms of pathogenesis of Bacillus anthracis.
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PMID:Anthrax toxin. 1159 78

Anthrax toxin consists of three nontoxic proteins that associate in binary or ternary combinations to form toxic complexes at the surface of mammalian cells. One of these proteins, protective antigen (PA), transports the other two, edema factor (EF) and lethal factor (LF), to the cytosol. LF is a Zn2+-protease that cleaves certain MAP kinase kinases, leading to death of the host via a poorly defined sequence of events. EF, a calmodulin- and Ca2+-dependent adenylate cyclase, is responsible for the edema seen in the disease. Both enzymes are believed to benefit the bacteria by inhibiting cells of the host's innate immune system. Assembly of toxic complexes begins after PA binds to cellular receptors and is cleaved into two fragments by furin proteases. The smaller fragment dissociates, allowing the receptor-bound fragment, PA63 (63 kDa), to self-associate and form a ring-shaped, heptameric pore precursor (prepore). The prepore binds up to three molecules of EF and/or LF, and the resulting complexes are endocytosed and trafficked to an acidic compartment. There, the prepore converts to a transmembrane pore, mediating translocation of EF and LF to the cytosol. Recent studies have revealed (a) the identity of receptors; (b) crystallographic structures of the three toxin proteins and the heptameric PA63 prepore; and (c) information about toxin assembly, entry, and action within the cytosol. Knowledge of the structure and mode of action of the toxin has unveiled potential applications in medicine, including approaches to treating anthrax infections.
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PMID:Anthrax toxin. 1457 May 63

Bacillus anthracis, the etiological agent of anthrax, secretes three polypeptides that assemble into toxic complexes on the cell surfaces of the host it infects. One of these polypeptides, protective antigen (PA), binds to the integrin-like domains of ubiquitously expressed membrane proteins of mammalian cells. PA is then cleaved by membrane endoproteases of the furin family. Cleaved PA molecules assemble into heptamers, which can then associate with the two other secreted polypeptides: edema factor (EF) and/or lethal factor (LF). The heptamers of PA are relocalized to lipid rafts where they are quickly endocytosed and routed to an acidic compartment. The low pH triggers a conformational change in the heptamers, resulting in the formation of cation-specific channels and the translocation of EF/LF. EF is a calcium- and calmodulin-dependent adenylate cyclase that dramatically raises the intracellular concentration of cyclic adenosine monophosphate (cAMP). LF is a zinc-dependent endoprotease that cleaves the amino terminus of mitogen-activated protein kinase kinases (Meks). Cleaved Meks cannot bind to their substrates and have reduced kinase activity, resulting in alterations of the signaling pathways they govern. The structures of PA, PA heptamer, EF, and LF have been solved and much is now known about the molecular details of the intoxication mechanism. The in vivo action of the toxins, on the other hand, is still poorly understood and hotly debated. A better understanding of the toxins will help in the design of much-needed anti-toxin drugs and the development of new toxin-based medical applications.
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PMID:Anthrax toxins. 1554 6

Several molecular models of glycoprotein hormone receptor activation have been proposed. It has been suggested that ligand binding to the ectodomain (ECD) leads to major changes in intramolecular interactions between the ECD and the transmembrane domain. We studied these intramolecular modifications by generating a recombinant LH/CG receptor (LHR) bearing an intramolecular cleavage site. We did this by inserting a furin site at position 316 in the hinge region of the ECD (LHR_Fur316). Affinity for human chorionic gonadotropin (hCG) and cAMP production upon hCG stimulation was identical to those of wild-type LHR. Western blot analysis showed that the LHR_Fur316 receptor was cleaved into two subunits linked by disulfide bridges. Chemical shedding of the ECD from the transmembrane domain did not increase basal adenylate cyclase activity, indicating that the first 294 residues did not act as an inverse agonist. The truncated LHR_316 was still activated by hCG but with an EC50 higher than that for the wild-type receptor. Zero length cross-linking was used to study intramolecular interactions between the two domains of LHR_Fur316. Cross-linking efficiency was similar for the basal and activated states, which indicated that the two domains interacted closely in the basal state, and this tight interaction persisted during activation. Our data suggest that activation of the LHR results from subtle modifications of intramolecular interactions between the two domains and low-affinity binding of hCG to the extracellular loops or residues preceding the first transmembrane segment.
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PMID:Zero-length cross-linking reveals that tight interactions between the extracellular and transmembrane domains of the luteinizing hormone receptor persist during receptor activation. 1587 56

Endothelin-1 (ET-1) is an extremely potent vasoconstrictor peptide originally isolated from endothelial cells. Its synthesis, mainly regulated at the gene transcription level, involves processing of a precursor by a furin-type proprotein convertase to an inactive intermediate, big ET-1. The latter peptide can then be cleaved directly by an endothelin-converting enzyme (ECE) into ET-1 or reach the active metabolite through a two-step process involving chymase hydrolyzing big ET-1 to ET-1 (1-31), itself needing conversion to ET-1 by neprilysin (NEP) to exert physiological activity. ET-1 signals through two G protein-coupled receptors, endothelin receptor A (ETA) and endothelin receptor B (ETB). Both receptors induce an increase in intracellular Ca(2+), mainly from the extracellular space through voltage-independent mechanisms, the receptor-operated channels and store-operated channels. ET-1 also induces signaling through epidermal growth factor receptor transactivation, oxidative stress induction, rho-kinase, and the activation (ETA) or inhibition (ETB) of the adenylate cyclase/cyclic adenosine monophosphate pathway. Arterial vasoconstriction is mediated mainly by the ETA receptor. ET-1, via endothelium-located ETB, relaxes arteries or constricts vessels following activation of the same receptor type on the smooth muscle, where it can interact with ETA. In addition, ETB-dependent vasoconstriction seems more prominent in the venous vasculature. A better understanding of how ET-1 is synthesized and how ETA and ETB receptors interact could help design better pharmacological agents in the treatment of cardiovascular diseases where targeting the ET-1 system is indicated.
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PMID:Endothelin-1: Biosynthesis, Signaling and Vasoreactivity. 2745 Oct 97


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