Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P08908 (5-HT1A)
 5,574 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We report the cloning and expression of a novel 5-HT receptor gene from human genomic DNA. This clone, HGCR1, contains an apparently intronless open reading frame of 390 amino acids with the seven hydrophobic regions, typical of G-protein coupled receptors. The deduced amino acid sequence of HGCR1 is 39%, 55% and 87% identical to that for the human 5-HT1A, the human 5-HT1D and the rat 5-HT1B receptor, respectively. [3H]5-HT binding to transfected COS-7 cell membranes yields a pharmacological profile similar to that of 5-HT1B receptor. Thus these findings indicate the presence of 5-HT1B-type receptor in the human.
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PMID:Cloning and expression of the human 5-HT1B-type receptor gene. 161 Mar 47

The signal transduction pathways of the cloned human 5-HT1A receptor have been examined in two mammalian cell lines transiently (COS-7) or permanently (HeLa) expressing this receptor gene. In both systems, 5-hydroxytryptamine (5-HT, serotonin) mediated a marked inhibition of beta 2-adrenergic agonist-stimulated (80% inhibition in COS-7 cells) or forskolin-stimulated cAMP formation (up to 90% inhibition in HeLa cells). This serotonin effect (EC50 = 20 nM) could be competitively antagonized by metitepine and spiperone (Ki = 81 and 31 nM, respectively) and could also be blocked by pretreatment of cells with pertussis toxin. In both cell types, 5-HT failed to stimulate adenylyl cyclase through the expressed receptors. In HeLa cells, 5-HT also stimulated phospholipase C (approximately 40-75% stimulation of formation of inositol phosphates). Again, this effect was inhibited by metitepine. However, the EC50 of 5-HT was considerably higher (approximately 3.2 microM) than that found for inhibition of adenylyl cyclase. Both pathways were demonstrated to be similarly affected by pertussis toxin. These findings indicate that like the M2 and M3 muscarinic cholinergic receptors, the 5-HT1A receptor can couple to multiple transduction pathways with varying efficiencies via pertussis toxin-sensitive G-proteins. The lack of stimulation of cAMP formation by this 5-HT1A receptor may suggest the existence of another pharmacologically closely related receptor.
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PMID:Effector coupling mechanisms of the cloned 5-HT1A receptor. 254 39

alpha2-Adrenergic receptors (alpha 2AR) functionally couple not only to Gi but also to Gs. We investigated the amino-terminal portion of the third intracellular loop of the human alpha 2AAR (alpha 2C10) for potential Gs coupling domains using site-directed mutagenesis and recombinant expression in several different cell types. A deletion mutant and four chimeric receptors consisting of the alpha 2AAR with the analogous sequence from the 5-HT1A receptor (a Gi-coupled receptor) and the beta 2AR (a Gs-coupled receptor) were expressed in Chinese hamster ovary cells, Chinese hamster fibroblasts, or COS-7 cells and examined for their ability to mediate stimulation or inhibition of membrane adenylyl cyclase activity or whole cell cAMP accumulation. In stably expressing Chinese hamster ovary cells, deletion of amino acids 221-231, which are in close proximity to the fifth transmembrane domain, eliminated alpha 2C10-mediated stimulation of adenylyl cyclase activity, while alpha 2C10-mediated inhibition was only moderately affected. This suggested that this region is important for Gs coupling, prompting construction of the chimeric receptor mutants. Substitution of amino acids 218-235 with 5-HT1A receptor sequence entirely ablated agonist-promoted Gs coupling, as compared with a 338 +/- 29% stimulation of adenylyl cyclase activity observed with the wild-type alpha 2C10. In contrast, Gi coupling for this mutant remained fully intact (57 +/- 2% versus 52 +/- 1% inhibition for wild-type alpha 2C10). Similar substitution with beta 2AR sequence had no effect on Gi coupling but did reduce Gs coupling. Two additional mutated alpha 2C10 containing smaller substitutions of the amino-terminal region with 5-HT1A receptor sequence at residues 218-228 or 229-235 were then studied. While Gi coupling remained intact with both mutants, Gs coupling was ablated in the former but not the latter mutant receptor. Similar results were obtained using transfected Chinese hamster fibroblasts (which exclusively display alpha 2AR-Gi coupling) and COS-7 cells (which exclusively display alpha 2AR-Gs coupling). Thus, a critical determinant for Gs coupling is contained within 11 amino acids (218-228) of the amino-terminal region of the third intracellular loop localized directly adjacent to the fifth transmembrane domain. Taken together, these studies demonstrate the presence of a discrete structural determinant for agonist-promoted alpha 2AR-Gs coupling, which is distinct and separable from the structural requirements for alpha 2AR-Gi coupling.
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PMID:Identification of a Gs coupling domain in the amino terminus of the third intracellular loop of the alpha 2A-adrenergic receptor. Evidence for distinct structural determinants that confer Gs versus Gi coupling. 755 92

We have used the polymerase chain reaction technique to selectively amplify a guanine nucleotide-binding protein-coupled receptor cDNA sequence from rat striatal mRNA that exhibits high homology to previously cloned serotonin receptors. Sequencing of a full length clone isolated from a rat striatal cDNA library revealed an open reading frame of 1311 base pairs, encoding a 437-residue protein with seven hydrophobic regions. Within these hydrophobic regions, this receptor was found to be 41-36% identical to the following serotonin [5-hydroxytryptamine (5-HT)] receptors: 5-HT2 > 5-HT1D > 5-HT1C > 5-HT1B > 5-HT1A > 5-HT1E. Northern blots revealed a approximately 4.2-kilobase transcript localized in various brain regions, with the following rank order of abundance: striatum >> olfactory tubercle > cerebral cortex > hippocampus. Expression of this clone in COS-7 cells resulted in the appearance of high affinity, saturable binding of (+)-[2-125I] iodolysergic acid diethylamide ([125I]LSD) with a Kd of 1.26 nM. Among endogenous biogenic amines, only 5-HT completely inhibited [125I]LSD binding (Ki = 150 nM). The inhibition of [125I]LSD binding by other serotonergic agonists and antagonists revealed a pharmacological profile that does not correlate with that of any previously described serotonin receptor subtype. In addition, this receptor exhibits high affinity for a number of tricyclic antipsychotic and antidepressant drugs, including clozapine, amoxipine, and amitriptyline. In HEK-293 cells stably transfected with this receptor, serotonin elicits a potent stimulation of adenylyl cyclase activity, which is blocked by antipsychotic and antidepressant drugs. The distinct structural and pharmacological properties of this receptor site indicate that it represents a completely novel subtype of serotonin receptor. Based on its affinity for tricyclic psychotropic drugs and its localization to limbic and cortical regions of the brain, it is likely that this receptor may play a role in several neuropsychiatric disorders that involve serotonergic systems.
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PMID:Cloning and expression of a novel serotonin receptor with high affinity for tricyclic psychotropic drugs. 768 Jul 51

We report two serotonin (5-hydroxytryptamine, 5-HT) receptors, MR22 and REC17, that belong to the G-protein-associated receptor superfamily. MR22 and REC17 are 371 and 357 amino acids long, respectively, as deduced from nucleotide sequence and share 68% mutual amino acid identity and 30-35% identity with known catecholamine and 5-HT receptors. Saturable binding of 125I-labeled (+)-lysergic acid diethylamide to transiently expressed MR22 in COS-M6 cells was inhibited by ergotamine > methiothepin > 5-carboxamidotryptamine > 5-HT. For REC17, the rank of potency was ergotamine > 5-carboxamidotryptamine > methiothepin > methysergide > 5-HT. Both were insensitive to 5-HT1A, 5-HT1D or 5-HT2 serotonergic ligands [8-hydroxy-2-(di-n-propylamino)tetralin, sumatriptan, and 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane]. The mRNAs encoding MR22 were detected in the CA1 region of hippocampus, the medial habenula, and raphe nuclei. In contrast, mRNAs encoding REC17 were found throughout the rat central nervous system. We propose that REC17 and MR22, designated as 5-HT5 alpha and 5-HT5 beta, represent a distinct subfamily of 5-HT receptors.
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PMID:Two members of a distinct subfamily of 5-hydroxytryptamine receptors differentially expressed in rat brain. 768 2

The serotonin 5-HT1B and 5-HT1A receptors bind certain beta-adrenergic antagonists, such as propranolol and pindolol, with high affinity. Other 5-HT1 receptors that display very low affinity for beta-adrenergic antagonists, have either a threonine (T) (5-HT1D alpha, 5-HT1D beta and 5-HT1E) or an alanine (A) (5-HT1F) residue in the homologous position in the seventh transmembrane domain. In the case of the human 5-HT1D beta receptor, replacement of this T with asparagine (N), dramatically increases its ability to bind beta-adrenergic antagonists. To assess whether other 5-HT1 receptors would behave similarly, we have used site-directed mutagenesis to replace the T or A in 5-HT1D alpha, 5-HT1E and 5-HT1F receptors with N. Both the wild-type and mutant genes were expressed transiently in COS-7 cells and radioligand binding studies were performed by using [3H]5-HT and [125I]iodocyanopindolol. Using [3H]5-HT, we found that the affinities of all the mutant receptors for propranolol and pindolol were significantly increased by 100-1000 fold, 5-HT1D alpha and 5-HT1F receptors showing the highest and the 5-HT1E receptor displaying the lowest affinity. On the other hand, the affinities for 5-HT were essentially unchanged as compared to the wild-type receptors. All mutant receptors bound [125I]iodocyanopindolol with high affinity, KD values ranging between 0.04 nM (mutant 5-HT1D alpha) and 0.57 nM (mutant 5-HT1E), whereas the wild-type receptors failed to show any specific binding with this radioligand in the same concentration range used for the mutant receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A single point mutation increases the affinity of serotonin 5-HT1D alpha, 5-HT1D beta, 5-HT1E and 5-HT1F receptors for beta-adrenergic antagonists. 798 76

The functional significance of the conserved amino acids within transmembrane regions II and VII of the human 5-hydroxytryptamine (5-HT)1A receptor was analyzed by oligonucleotide-directed mutagenesis followed by transient expression of the mutated receptor genes in COS-1 cells. The substitution of a conserved asparagine at position 396 (transmembrane region VII) with either alanine, phenylalanine, or valine resulted in a receptor that did not bind the 5-HT1A agonist 8-hydroxy-2-(di-n-[3H]propylamino)tetralin. In contrast, replacement of Asn396 with glutamine did not affect agonist binding. In addition, serine residues at positions 391 and 393 (transmembrane domain VII) were changed to alanine. Changing the less conserved Ser391 to alanine had no effect on ligand binding. However, replacement of the conserved Ser393 with alanine reduced ligand binding by 86%. Replacement of a conserved aspartate at position 82 (transmembrane region II) with alanine also produced a receptor without detectable agonist binding. Protein immunoblotting detected receptor protein of approximately 51 kDa in both wild-type and mutant receptor-expressing cells, indicating that these mutations probably did not affect expression or processing of the protein. Importantly, the sequence of the human 5-HT1A receptor described in this paper differs from the published sequence [Nature (Lond.) 329:75-79 (1987)] in transmembrane region IV. The present sequence encodes a protein of 422 amino acids, instead of the 421-amino acid protein that has been described previously [Nature (Lond.) 329:75-79 (1987)], and has a change in the sequence in transmembrane region IV from ... RPRAL ... to ... RRAAA ..., which corresponds to the published sequence [J. Biol. Chem. 265:5825-5832 (1990)] of the rat 5-HT1A receptor. Moreover, conversion of the transmembrane region IV sequence of the present clone to that of the published sequence by site-directed mutagenesis abolished ligand binding to the receptor.
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PMID:Identification of residues important for ligand binding to the human 5-hydroxytryptamine1A serotonin receptor. 847 30

The cDNA from a schizophrenic patient heterozygous for a mutation of the 5-HT1A receptor gene was used to clone the variant and wild-type DNA into a eukaryotic expression vector. The mutation was characterized by a base pair substitution (A --> G) at the first position of codon 28, leading to an Ile --> Val amino acid exchange. COS-7 cells were transfected with the cDNA of either the wild type or the variant 5-HT1A receptor. The potencies of the 5-HT1A receptor agonists 8-hydroxy-2-(di-n-propylamino) tetraline (8-OH-DPAT), 5-HT and roxindole, and of the antagonists methiothepin and spiperone in inhibiting specific binding of [3H]8-OH-DPAT of the mutant and wild-type 5-HT1A receptor ligands concentration-dependently inhibited specific [3H]8-OH-DPAT binding to both the wild-type and the variant 5-HT1A receptor. The rank order of potency of the ligands in inhibiting [3H]8-OH-DPAT binding was identical at both receptors and was roxindole > 8-OH-DPAT > 5-HT> methiothepin > spiperone. This rank order is characteristic for 5-HT1A receptors. The negative logarithms of the concentrations required for 50% inhibition (pIC50 values) of the ligands at the mutant 5-HT receptor correlated highly significantly with those at the wildtype receptor (r = 0.995). It is concluded that the pharmacological profile of the mutant 5-HT1A receptor does not differ from that of the wild-type 5-HT1A receptor.
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PMID:Binding properties of the naturally occurring human 5-HT1A receptor variant with the Ile28Val substitution in the extracellular domain. 853 77

We have cloned by homology screening from a rat brain cDNA library a GIRK3-type (Kir 3.3) inwardly rectifying K+ channel subunit with high structural similarity to other subfamily members whose activity is thought to be controlled by receptor-stimulated G proteins. When heterologously expressed both in Xenopus oocytes and in mammalian COS-7 cells, rbGIRK3 subunits individually fail to form functional channels. In contrast, when coexpressed with other GIRK subunits, rbGIRK3 gives rise to prominent currents which are enhanced by the stimulation of coexpressed 5-HT1A receptors. In situ hybridizations show that of all GIRK subunits rbGIRK3 is most widely distributed and strongly expressed throughout the rat brain and thus may play an important role in central signal processing.
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PMID:Functional expression and cellular mRNA localization of a G protein-activated K+ inward rectifier isolated from rat brain. 867 Mar 6

Differential immunohistochemical labeling is often observed using different antibodies against the same protein. Two polyclonal antipeptide antibodies against the 5-HT1A receptor have been generated by our group. The S1A-170 (aa 170-186) and 258 (aa 258-274) are specific for sites in the second extracellular loop and third intracellular loop, respectively [E.C. Azmtia, I. Yu, H.M. Akbari, N. Kheck, P.M. Whitaker-Azmitia and D.R. Marshak, Antipeptide antibodies against the 5-HT1A receptor, J. Chem. Neuroanat., 5 (1992) 289-298]. Comparison of the labeling patterns of these two antibodies and other antipeptide antibodies against the 5-HT1A receptor revealed that although similar populations of cells were labeled, individual antibodies favor certain staining patterns. Immunocytochemistry and western blotting results of transfected cell lines and brain tissue revealed the following: (1) both the S1A-170 and S1A-258 are specific for the 5-HT1A receptor when used for immunocytochemistry in transfected HEK-293 and COS-1 cells; (2) when expressed in cultured cell lines, the 5-HT1A receptor is differentially glycosylated dependent on cell type, and the S1A-258 is specific for only certain species on immunoblots; and (3) the S1A-258 and L5B7 [M. Riad, S. El Mestikawy, D. Derge, H. Gozlan, and M. Hamon, Visualization and quantification of central 5-HT1A receptors with specific antibodies, Neurochem. Int., 4 (1991) 413-423] label common bands at 40 and 70 kDa on immunoblots of hippocampal proteins, but show opposite staining intensities. These results provide evidence for the immunocytochemical specificity of both the S1A-170 and S1A-258 and suggest that the discrepancies noted in immunohistochemistry may be due in part to different molecular conformations.
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PMID:Molecular characterization of antipeptide antibodies against the 5-HT1A receptor: evidence for state-dependent antibody binding. 940 44


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