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Title: Cloning, sequencing, and expression of the gene coding for the human platelet. cap alpha. /sub 2/-adrenergic receptor

Abstract

The gene for the human platelet ..cap alpha../sub 2/-adrenergic receptor has been cloned with oligonucleotides corresponding to the partial amino acid sequence of the purified receptor. The identity of this gene has been confirmed by the binding of ..cap alpha../sub 2/-adrenergic ligands to the cloned receptor expressed in Xenopus laevis oocytes. The deduced amino acid sequence is most similar to the recently cloned human ..beta../sub 2/- and ..beta../sub 1/-adrenergic receptors; however, similarities to the muscarinic cholinergic receptors are also evident. Two related genes have been identified by low stringency Southern blot analysis. These genes may represent additional ..cap alpha../sub 2/-adrenergic receptor subtypes.

Authors:
; ; ; ; ; ; ;
Publication Date:
Research Org.:
Duke Univ. Medical Center, Durham, NC (USA)
OSTI Identifier:
6805416
Resource Type:
Journal Article
Resource Relation:
Journal Name: Science (Washington, D.C.); (United States); Journal Volume: 238
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; GENES; CLONING; DNA SEQUENCING; RECEPTORS; AMINO ACID SEQUENCE; SYMPATHOMIMETICS; GENE REGULATION; BLOOD PLATELETS; FROGS; LIGANDS; MAN; OOCYTES; AMPHIBIANS; ANIMALS; AQUATIC ORGANISMS; AUTONOMIC NERVOUS SYSTEM AGENTS; BIOLOGICAL MATERIALS; BLOOD; BLOOD CELLS; BODY FLUIDS; DRUGS; GERM CELLS; MAMMALS; MATERIALS; MEMBRANE PROTEINS; MOLECULAR STRUCTURE; ORGANIC COMPOUNDS; PRIMATES; PROTEINS; STRUCTURAL CHEMICAL ANALYSIS; VERTEBRATES 550200* -- Biochemistry

Citation Formats

Kobilka, B.K., Matsui, H., Kobilka, T.S., Yang-Feng, T.L., Francke, U., Caron, M.G., Lefkowitz, R.J., and Regan, J.W. Cloning, sequencing, and expression of the gene coding for the human platelet. cap alpha. /sub 2/-adrenergic receptor. United States: N. p., 1987. Web. doi:10.1126/science.2823383.
Kobilka, B.K., Matsui, H., Kobilka, T.S., Yang-Feng, T.L., Francke, U., Caron, M.G., Lefkowitz, R.J., & Regan, J.W. Cloning, sequencing, and expression of the gene coding for the human platelet. cap alpha. /sub 2/-adrenergic receptor. United States. doi:10.1126/science.2823383.
Kobilka, B.K., Matsui, H., Kobilka, T.S., Yang-Feng, T.L., Francke, U., Caron, M.G., Lefkowitz, R.J., and Regan, J.W. 1987. "Cloning, sequencing, and expression of the gene coding for the human platelet. cap alpha. /sub 2/-adrenergic receptor". United States. doi:10.1126/science.2823383.
@article{osti_6805416,
title = {Cloning, sequencing, and expression of the gene coding for the human platelet. cap alpha. /sub 2/-adrenergic receptor},
author = {Kobilka, B.K. and Matsui, H. and Kobilka, T.S. and Yang-Feng, T.L. and Francke, U. and Caron, M.G. and Lefkowitz, R.J. and Regan, J.W.},
abstractNote = {The gene for the human platelet ..cap alpha../sub 2/-adrenergic receptor has been cloned with oligonucleotides corresponding to the partial amino acid sequence of the purified receptor. The identity of this gene has been confirmed by the binding of ..cap alpha../sub 2/-adrenergic ligands to the cloned receptor expressed in Xenopus laevis oocytes. The deduced amino acid sequence is most similar to the recently cloned human ..beta../sub 2/- and ..beta../sub 1/-adrenergic receptors; however, similarities to the muscarinic cholinergic receptors are also evident. Two related genes have been identified by low stringency Southern blot analysis. These genes may represent additional ..cap alpha../sub 2/-adrenergic receptor subtypes.},
doi = {10.1126/science.2823383},
journal = {Science (Washington, D.C.); (United States)},
number = ,
volume = 238,
place = {United States},
year = 1987,
month =
}
  • Pharmacologic, biochemical, and genetic analyses have demonstrated the existence of multiple {alpha}{sub 2}-adrenergic receptor ({alpha}{sub 2}AR) subtypes. The authors have cloned a human {alpha}{sub 2}AR by using the polymerase chain reaction with oligonucleotide primers homologous to conserved regions of the previously cloned {alpha}{sub 2}ARs, the genes for which are located on human chromosomes 4 (C4) and 10 (C10). The deduced amino acid sequence encodes a protein of 450 amino acids whose putative topology is similar to that of the family of guanine nucleotide-binding protein-coupled receptors, but whose structure most closely resembles that of the {alpha}{sub 2}ARs. Competition curve analysis ofmore » the binding properties of the receptor expressed in COS-7 cells with a variety of adrenergic ligands demonstrates a unique {alpha}{sub 2}AR pharmacology. Hybridization with somatic cell hybrids shows that the gene for this receptor is located on chromosome 2. Northern blot analysis of various rat tissues shows expression in liver and kidney. The unique pharmacology and tissue localization of this receptor suggest that this is an {alpha}{sub 2}AR subtype not previously identified by classical pharmacological or ligand binding approaches.« less
  • The complete nucleotide sequence of a cDNA encoding the human platelet-derived growth factor (PDGF) receptor is presented. The cDNA contains an open reading frame that codes for a protein of 1106 amino acids. Comparison to the mouse PDGF receptor reveals an overall amino acid sequence identity of 86%. This sequence identity rises to 98% in the cytoplasmic split tyrosine kinase domain. RNA blot hybridization analysis of poly(A){sup +} RNA from human dermal fibroblasts detects a major and a minor transcript using the cDNA as a probe. Baby hamster kidney cells, transfected with an expression vector containing the receptor cDNA, expressmore » an {approx} 190-kDa cell surface protein that is recognized by an anti-human PDGF receptor antibody. The recombinant PDGF receptor is functional in the transfected baby hamster kidney cells as demonstrated by ligand-induced phosphorylation of the receptor. Binding properties of the recombinant PDGF receptor were also assessed with pure preparations of BB and AB isoforms of PDGF. Unlike human dermal fibroblasts, which bind both isoforms with high affinity, the transfected baby hamster kidney cells bind only the BB isoform of PDGF with high affinity. This observation is consistent with the existence of more than one PDGF receptor class.« less
  • We report the molecular cloning of the human gene (symbol LRPAP1) coding for the {alpha}{sub 2}-macroglobulin receptor-associated protein (A2MRAP), as well as the gene coding for the 44-kDa heparin-binding protein (HBP-44), its murine counterpart. For both, genomic cosmid clones were isolated, and for the human gene a bacteriophage P1 clone containing the entire A2MRAP gene was also retrieved. The genes were characterized after subcloning: in both species, the known coding part of the cDNA is encoded by eight exons, and the position of the boundaries of the exons was conserved. The human LRPAP1 locus was assigned to chromosome 4 bymore » PCR of human-hamster hybrid cell lines and by fluorescence in situ hybridization to band 4p16.3. This maps closely to the variable constitutional deletions of the short arm of chromosome 4, observed cytogenetically in patients with the Wolf-Hirschhorn syndrome. Metaphase spreads of two such patients were analyzed by fluorescence in situ hybridization with an LRPAP1 genomic probe. The first patient, with karyotype 46,XY,del4(p14-p16.1), had retained both copies of the LRPAP1 gene. In contrast, the other patient, with karyotype 46,XY,del4(p15.3-pter), displayed no signal for LRPAP1 on the deleted chromosome. 44 refs., 3 figs., 1 tab.« less
  • An /alpha//sub 2/-adrenergic receptor subtype has been cloned from a human kidney cDNA library using the gene for the human platelet /alpha//sub 2/-adrenergic receptor as a probe. The deduced amino acid sequence resembles the human platelet /alpha//sub 2/-adrenergic receptor and is consistent with the structure of other members of he family of guanine nucleotide-binding protein-coupled receptors. The cDNA was expressed in a mammalian cell line (COS-7), and the /alpha//sub 2/-adrenergic ligand (/sup 3/H)rauwolscine was bound. Competition curve analysis with a variety of adrenergic ligands suggests that this cDNA clone represents the /alpha//sub 2/B-adrenergic receptor. The gene for this receptor ismore » on human chromosome 4, whereas the gene for the human platelet /alpha//sub 2/-adrenergic receptor (/alpha//sub 2/A) lies on chromosome 10. This ability to express the receptor in mammalian cells, free of other adrenergic receptor subtypes, should help in developing more selective /alpha/-adrenergic ligands.« less
  • The authors have isolated and sequenced a cDNA encoding the human ..beta../sub 2/-adrenergic receptor. The deduced amino acid sequence (413 residues) is that of a protein containing seven clusters of hydrophobic amino acids suggestive of membrane-spanning domains. While the protein is 87% identical overall with the previously cloned hamster ..beta../sub 2/-adrenergic receptor, the most highly conserved regions are the putative transmembrane helices (95% identical) and cytoplasmic loops (93% identical), suggesting that these regions of the molecule harbor important functional domains. Several of the transmembrane helices also share lesser degrees of identity with comparable regions of select members of the opsinmore » family of visual pigments. They have localized the gene for the ..beta../sub 2/-adrenergic receptor to q31-q32 on chromosome 5. This is the same position recently determined for the gene encoding the receptor for platelet-derived growth factor and is adjacent to that for the FMS protooncogene, which encodes the receptor for the macrophage colony-stimulating factor.« less