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 mechanism of homologous regulation of MSH receptor binding and receptor-mediated adenylate cyclase activation in three human and two mouse melanoma cell lines. Pretreatment with alpha-MSH resulted in a time- and dose-dependent up-regulation of MSH receptors in human D10 and 205 melanoma cells whereas in human HBL and in mouse B16-F1 and Cloudman S91 cells alpha-MSH induced receptor down-regulation. Up-regulation of receptors was maximal after a 24-h incubation period and an alpha-MSH concentration of 100 nM (EC50 = 2.4 nM). The increase in alpha-MSH binding was independent of adenylate cyclase activation and protein synthesis and appeared to be caused by recruitment of spare receptors. The structural requirements of the peptide for triggering this process differed from those found in receptor-binding analyses. Receptor down-regulation was maximal after 12 h and hence more rapid than up-regulation. In B16-F1 cells, 10 nM alpha-MSH caused the disappearance of 85-90% of the MSH receptors, the EC50 of 0.23 nM lying exactly between that for alpha-MSH-induced melanogenesis (0.027 nM) and the dissociation constant of receptor binding (1.31 nM). Down-regulation in B16-F1 cells appears to be the consequence of receptor internalization following MSH binding and seems to be initiated during an early step in MSH signalling, preceding the activation of adenylate cyclase and the cAMP signal. Receptor up- and down-regulation were not accompanied by an alteration in affinity to alpha-MSH, as demonstrated by Scatchard analysis of the binding curves.
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PMID:Homologous regulation of the MSH receptor in melanoma cells. 838 55

The regulation of adenylate cyclase activity by adrenocorticotropin/alpha-melanocyte-stimulating hormone (ACTH/MSH)-like peptides was investigated in rat brain slices using a superfusion method. Adenylate cyclase activity was concentration-dependently increased by ACTH-(1-24), alpha-MSH (EC50 values 16 and 6 nM, respectively), and [Nle4,D-Phe7]alpha-MSH (EC50 value 1.6 nM), in the presence of forskolin (1 microM, optimal concentration). 1-9-Dideoxyforskolin did not augment the response of adenylate cyclase to ACTH-(1-24). Various peptide fragments were tested for their ability to enhance [3H]cyclic AMP production. [Nle4,D-Phe7]alpha-MSH increased [3H]cyclic AMP formation with a maximal effect of 30% and was more potent than ACTH-(1-24), ACTH-(1-16)-NH2, alpha-MSH, ACTH-(1-13)-NH2, [MetO4]alpha-MSH, [MetO2(4),D-Lys8,Phe9]ACTH-(4-9), ACTH-(7-16)-NH2, ACTH-(1-10), and ACTH-(11-24), in order of potency. This structure-activity relationship resembles that found for the previously described peptide-induced display of excessive grooming. ACTH-(1-24) stimulated adenylate cyclase activity in both striatal (maximal effect, approximately 20%) and septal slices (maximal effect, approximately 40%), but not in hippocampal or cortical slices. Lesioning of the dopaminergic projections to the striatum did not result in a diminished effect of [Nle4,D-Phe7]alpha-MSH on [3H]cyclic AMP accumulation, which indicates that the ACTH/MSH receptor-stimulated adenylate cyclase is not located on striatal dopaminergic terminals. ACTH-(1-24) did not affect the dopamine D1 or D2 receptor-mediated modulation of adenylate cyclase activity.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Adrenocorticotropin/alpha-melanocyte-stimulating hormone (ACTH/MSH)-like peptides modulate adenylate cyclase activity in rat brain slices: evidence for an ACTH/MSH receptor-coupled mechanism. 838 34

Melanocortins, melanocyte-stimulating hormones (MSH) and adrenocorticotropic hormone (ACTH) are homologous natural peptides derived from pro-opiomelanocortin (POMC). Recent breakthroughs in melanocortin receptor (MCR) biology are relevant to neuroimmunomodulation because melanocortins are known to modulate fever, inflammation and immunity, by acting both on peripheral targets and within the brain. During fever, endogenous melanocortins exert antipyretic effects by acting on MCR located within the brain, suggesting a protective counterregulatory role of the central melanocortin system. MCR are also found in melanocytic cells and adrenal cortical cells, the classical targets for alpha-MSH and ACTH, respectively, in myelogenous and lymphoid tissues, and in various endocrine and exocrine glands, adipocytes, and in autonomic ganglia. In the CNS, MCR are prominently distributed in close proximity to the terminal fields of melanocortinergic neurons that innervate neuroendocrine and autonomic motor nuclei as well as other subcortical brain regions important in neuroendocrine and autonomic regulation, sensory processing and various aspects of behavior. Furthermore, the presence of MCR in circumventricular organs of the brain provides direct access of systemic melanocortin hormones to central MCR. Together, these attributes provide an anatomical basis for bidirectional MCR-mediated communication between brain and periphery. A group of five G-protein-associated MCR subtypes, each of which is positively coupled to adenylate cyclase, has been identified. Among these, the adrenal ACTH receptor (MC2-R) is selectively activated by ACTH. In contrast, the other MCR subtypes (MC1-R, MC3-R, MC4-R, MC5-R) recognize a common group of ligands that includes various forms of MSH as well as ACTH; nevertheless they do exhibit important differences in ligand selectivity. MCR concentrations and MCR mRNA levels are influenced by availability of cognate ligands, by drugs, and by pathological stimuli. Two types of endogenous MCR antagonist proteins have been discovered: agouti protein and the corticostatins. Agouti protein dramatically alters coat color in mammals by antagonizing melanocytic MC1-R. Moreover, spontaneous dominant mutations of the agouti gene in several strains of mice lead to its ubiquitous overexpression and produces not only yellow coat color, but also obesity and insulin resistance, perhaps as a result of its antagonism of other MCR subtypes. The recent emergence of synthetic MCR antagonists, and the feasibility of molecular approaches for targeted inactivation of individual MCR subtypes, should facilitate elucidation of the roles and mechanisms of neuroimmunomodulation by endogenous melanocortins, and the determination of whether selective pharmacological targeting of MCR may ultimately have therapeutic utility.
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PMID:Receptor biology of the melanocortins, a family of neuroimmunomodulatory peptides. 921 48

An analogue of human melanin-concentrating hormone (MCH) suitable for radioiodination was designed in which Tyr13 and Val19 of the natural peptide were replaced by phenylalanyl and tyrosyl residues: [Phe13, Tyr19]-MCH. The peptide was synthesized by the continuous-flow solid-phase methodology using Fmoc-strategy and polyhipe PA 500 and PEG-PS resins. The linear MCH peptides with either acetamidomethyl-protected or free cysteinyl residues were purified to homogeneity and cyclized by iodine oxidation, yielding the final product with the correct molecular weight of 2434.61. Radioiodination of the C-terminal tyrosine was carried out enzymatically using solid-phase bound glucose oxidase/lactoperoxidase, followed by purification on a reversed-phase mini-column and by high-pressure liquid chromatography. The resulting [125I]-[Phe13, Tyr19]-MCH tracer was the first radiolabelled MCH peptide suitable for radioreceptor assay: saturation binding analysis using mouse G4F-7 melanoma cells demonstrated the presence of 1090 MCH receptors per cell. The dissociation constant (KD) was 1.18 x 10(-10) M, indicating high-affinity MCH receptors on these cells. MCH receptors were also found in other cell lines such as mouse B16-F1 and G4F and human RE melanoma cells as well as in PC12 and COS-7 cells. Competition binding analyses with a number of other peptides such as alpha-MSH, neuropeptide Y, substance P and pituitary adenylate cyclase activating peptide, demonstrated that the binding to the MCH receptor is specific. Atrial natriuretic factor was found to be a weak competitor of MCH, indicating topological similarities between MCH and ANF when interacting with MCH receptors.
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PMID:Synthesis and iodination of human (phenylalanine 13, tyrosine 19) melanin-concentrating hormone for radioreceptor assay. 922 84

Among the five members of the melanocortin receptor (MC-R) family, MC2 and MC5 are expressed in peripheral tissues. The receptor MC2 (ACTH receptor) almost exclusively expressed in the adrenal cortex whereas MC5-R is expressed in several organs including the adrenal cortex. Both receptors bind ACTH and activate adenylate cyclase. The aim of this work was to study the spatial distribution of MC5-R among the different zones of the bovine adrenal cortex and to analyze the regulation of its expression by its own ligands, ACTH and alpha-MSH and by angiotensin II (AII). Using semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and RNase protection assay, MC5-R was detected only in the glomerulosa zone whereas MC2-R was present in both glomerulosa and fasciculata zones of adult adrenal cortex. Treatments by ACTH, alpha-MSH, or AII increased the MC5-R mRNA level in glomerulosa cells by factors 7, 5, and 4.5, respectively. However, although potentially regulated by hormones, MC5-R is expressed at a level at least 100 times less than MC2-R, suggesting that MC5-R expression might only be at trace levels in grown adults, but could be much higher during embryogenesis.
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PMID:Expression and regulation of melanocortin receptor-5 (MC5-R) in the bovine adrenal cortex. 1068 56

The terminal differentiation of malignant melanoma cells is known to be induced by activating cAMP signaling pathway with alpha-MSH or cAMP analogues. However, sustained activation of cAMP signaling system that induces the differentiation of melanoma cells, also induces the desensitization of the pathway at the receptor level. Nevertheless, the adaptation of adenylate cyclase (AC) expression by sustained activation of cAMP signaling system has not been clearly understood. This study was performed to examine whether the sustained activation of cAMP system induce changes in the expression AC isoforms as an adaptation mechanism. Treatment of B16/F10 murine melanoma cells with 100 mM forskolin for 6 days resulted in differentiation, melanin accumulation and increased expression of tyrosine hydroxylase mRNA. In the forskolin-treated melanoma cells, change in expression of various AC isoform at the transcription level was detected by reverse-transcription polymerase chain reaction (RT-PCR). Expression of AC isoform mRNA: ACI, III, VI, VII, and IX increased to the level of 196-392% of the control whereas the level of ACII was decreased by 30%. The cAMP concentration was increased both in basal and alpha-MSH stimulated cells, but the AC activity was decreased in the forskolin treated cells. Thus, these results suggest that sustained activation of cAMP system induces differential expression of AC isoforms, which results in increase of cAMP accumulation.
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PMID:Multi-facet expressions of adenylate cyclase isoforms in B16-F10 melanoma cells differentiated by forskolin treatment. 1119 Feb 77

Membrane functions in tumorous cells are different from those in healthy cells. The aim of the present study was to investigate the changes in pituitary cell membrane functions and hormone secretion after tumor induction in vivo and in vitro. Prolactinomas were induced in vivo in female Wistar rats with estrone acetate. Normal anterior pituitaries and prolactinomas of female Wistar rats were dissociated enzymatically and mechanically, then cultured on collagen-treated plastic dishes. Some normal anterior pituitary cultures were treated with benz(c)acridines as tumorigenic agents in vitro. Intracellular 3',5'-cyclic-adenosine monophosphate (cAMP) levels were determined by a competitive binding technique, membrane fluidity was assayed by fluorescence anisotropy, and ATP-ase activities were estimated via ATP loss. The results indicated decreased membrane fluidity in tumorous cell cultures. However, in vitro benz(c)acridine treatment exerted more pronounced effects than those observed after in vivo estrone treatment. The ATP-ase activities were highly increased in benz(c)acridine-treated cells and in estrogen-induced prolactinoma cells, more strongly so in the former ones. The intracellular cAMP levels were higher than normal in both of them. The results concerning the ACTH, alpha-MSH, PRL and GH levels of normal and tumorous cell cultures were published in our previous study. Our findings show that the tumorous transformation of pituitary cells can cause significant changes in functional membrane parameters and hormone secretion. Decreased membrane fluidity was accompanied by an increased exocytosis (hormone release) and adenylate cyclase activity in tumorous cells.
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PMID:Functional membrane changes due to tumor induction in rat pituitary cell cultures. 1127 34

Among the melanocortins alpha-MSH is known to be involved in feeding behavior. These hormones mediate their effects through G protein-coupled receptors by stimulating adenylate cyclase. In this study, we have developed an in vitro expression model for human melanocortin 3 receptor (hMC3R) tagged at its C terminus with EGFP. The corresponding chimeric cDNA was stably expressed in HEK293 cells. The selected clones expressing the hMC3R-EGFP exhibited cell surface fluorescence and responded to NDP-MSH stimulation by producing cAMP in a dose-dependent manner (EC(50): 0.3 nM). Binding studies revealed a single class of binding sites with a K(D) of 2.24 nM. Moreover, Agouti-related protein was also demonstrated to be an antagonist of the hMC3R-EGFP. Thus, the hMC3R tagged with EGFP stably expressed in HEK293 cells, exhibiting the same characteristics than the wild-type hMC3R, is the only model of expression of this receptor allowing its direct localization inside living cells.
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PMID:Stable expression of human melanocortin 3 receptor fused to EGFP in the HEK293 cells. 1278 89

The melanocortin-4 receptor (MC4R) is a G-protein coupled receptor (GPCR) that is expressed in the central nervous system and has a role in regulating energy homeostasis and obesity. Up to a remarkable 6% of morbidly obese adults and children studied possess single nucleotide polymorphisms (SNPs) of the MC4R. Upon stimulation by agonist, the MC4R signals through the intracellular adenylate cyclase signal transduction pathway. Posttranslational modification of the pro-opiomelanocortin (POMC) gene transcript results in the generation of several endogenous melanocortin receptor agonists including alpha-, beta-, gamma-melanocyte stimulating hormones (MSH) and adrenocorticotropin (ACTH) ligands. The endogenous MC4R antagonist, agouti-related protein (AGRP), is expressed in the brain and is only one of two naturally occurring antagonists of GPCRs identified to date. Herein, we have generated 40 hMC4 polymorphic receptors and evaluated their cell surface expression by flow cytometry as well as pharmacologically characterized their functionality using the endogenous agonists alpha-MSH, beta-MSH, gamma2-MSH, ACTH(1-24), the antagonist hAGRP(87-132), and the synthetic agonists NDP-MSH and MTII. This is the first study in which polymorphic hMC4Rs have been pharmacologically characterized simultaneously with multiple endogenous ligands. Interestingly, at the N97D, L106P, and C271Y hMC4Rs beta-MSH was more potent than the other endogenous agonists alpha-MSH, gamma2-MSH, ACTH(1-24). The S58C and R165Q/W hMC4Rs possessed significantly reduced endogenous agonist potency (15- to 90-fold), but the synthetic ligands NDP-MSH and MTII possessed only 2-9-fold reduced potency as compared to the wild-type receptor, suggesting their potential as therapeutic ligands to treat individuals with these polymorphisms.
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PMID:Pharmacological characterization of 40 human melanocortin-4 receptor polymorphisms with the endogenous proopiomelanocortin-derived agonists and the agouti-related protein (AGRP) antagonist. 1675 16

Intensive efforts have been made to develop potent and selective ligands for certain human melanocortin receptors as possible treatments for obesity and sexual dysfunction due to the role of these receptors in feeding behavior, energy homeostasis, sexual function, etc. A number of novel alpha-MSH analogues were designed and synthesized primarily on the basis of our previous MTII NMR structure. In these peptide analogues, a disulfide or lactam bridge between residues at positions 5 and 8 was used as a conformational constraint to enhance the beta-turn spanning His6 and D-Phe7, while the pharmacophore group in Arg8 was mimicked via Nalpha-alkylation of residues 8 or 9 with the guanidinylbutyl group. Biological assays for binding affinities and adenylate cyclase activities for the hMC1R, hMC3R, hMC4R, and hMC5R showed that three analogues have good binding affinity for the hMC4R (0.7-4.1 nM), but have no binding affinity up to 10 microM at the other three melanocortin receptors. Interestingly, the three hMC4R selective analogues display only 50% binding efficiency, suggesting there is allosteric modulation of the melanocortin-4 receptor. These analogues were found to act as antagonists of the hMC4R. This result represents a discovery of very selective peptide-based antagonists for the hMC4R. The high selectivity may be due to the strong conformational constraint via ring contraction as compared to MTII, and the rigid conformation preferred by these new ligands allows them to recognize only the hMC4R, but not to activate the second messenger. The MTII NMR structure-based design thus not only examined the structural model of melanocortin ligands, but also yielded new biologically unique alpha-MSH analogues.
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PMID:Design, synthesis, and biological evaluation of new cyclic melanotropin peptide analogues selective for the human melanocortin-4 receptor. 1715 18


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