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. 1999 Oct 1;343(Pt 1):231–239.

Truncated isoforms inhibit [3H]prazosin binding and cellular trafficking of native human alpha1A-adrenoceptors.

F Cogé 1, S P Guenin 1, A Renouard-Try 1, H Rique 1, C Ouvry 1, N Fabry 1, P Beauverger 1, J P Nicolas 1, J P Galizzi 1, J A Boutin 1, E Canet 1
PMCID: PMC1220546  PMID: 10493934

Abstract

We have identified from human liver eight alpha(1A)-adrenoceptor (alpha(1A)-AR) splice variants that were also expressed in human heart, prostate and hippocampus. Three of these alpha(1A)-AR isoforms (alpha(1A-1)-AR, alpha(1A-2a)-AR and alpha(1A-3a)-AR) gave rise to receptors with seven transmembrane domains (7TMalpha(1A)-AR). The other five (alpha(1A-2b)-AR, alpha(1A-2c)-AR, alpha(1A-3c)-AR, alpha(1A-5)-AR and alpha(1A-6)-AR) led to truncated receptors lacking transmembrane domain VII (6TMalpha(1A)-AR). The 7TMalpha(1A)-AR isoforms transiently expressed in COS-7 cells bound [(3)H]prazosin with high affinity (K(d) 0.2 nM) and mediated a noradrenaline (norepinephrine)-induced increase in cytoplasmic free Ca(2+) concentration, whereas the 6TMalpha(1A)-AR isoforms were incapable of ligand binding and signal transduction. Immunocytochemical studies with N-terminal epitope-tagged alpha(1A)-AR isoforms showed that the 7TMalpha(1A)-AR isoforms were present both at the cell surface and in intracellular compartments, whereas the 6TMalpha(1A)-AR isoforms were exclusively localized within the cell. Interestingly, in co-transfected cells, each truncated alpha(1A)-AR isoform inhibited [(3)H]prazosin binding and cell-surface trafficking of the co-expressed 'original' 7TMalpha(1A-1)-AR. However, there was no modification of either the [(3)H]prazosin-binding affinity or the pharmacological properties of alpha(1A-1)-AR. Immunoblotting experiments revealed that co-expression of the alpha(1A-1)-AR with 6TMalpha(1A)-AR isoforms did not impair alpha(1A-1)-AR expression. Therefore the expression in human tissues of many truncated isoforms constitutes a new regulation pathway of biological properties of alpha(1A)-AR.

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Selected References

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  1. Autelitano D. J., Woodcock E. A. Selective activation of alpha1A-adrenergic receptors in neonatal cardiac myocytes is sufficient to cause hypertrophy and differential regulation of alpha1-adrenergic receptor subtype mRNAs. J Mol Cell Cardiol. 1998 Aug;30(8):1515–1523. doi: 10.1006/jmcc.1998.0717. [DOI] [PubMed] [Google Scholar]
  2. Berts A., Zhong H., Minneman K. P. No role for Ca++ or protein kinase C in alpha-1A adrenergic receptor activation of mitogen-activated protein kinase pathways in transfected PC12 cells. Mol Pharmacol. 1999 Feb;55(2):296–303. doi: 10.1124/mol.55.2.296. [DOI] [PubMed] [Google Scholar]
  3. Chang D. J., Chang T. K., Yamanishi S. S., Salazar F. H., Kosaka A. H., Khare R., Bhakta S., Jasper J. R., Shieh I. S., Lesnick J. D. Molecular cloning, genomic characterization and expression of novel human alpha1A-adrenoceptor isoforms. FEBS Lett. 1998 Jan 30;422(2):279–283. doi: 10.1016/s0014-5793(98)00024-6. [DOI] [PubMed] [Google Scholar]
  4. Daunt D. A., Hurt C., Hein L., Kallio J., Feng F., Kobilka B. K. Subtype-specific intracellular trafficking of alpha2-adrenergic receptors. Mol Pharmacol. 1997 May;51(5):711–720. doi: 10.1124/mol.51.5.711. [DOI] [PubMed] [Google Scholar]
  5. Eason M. G., Liggett S. B. Subtype-selective desensitization of alpha 2-adrenergic receptors. Different mechanisms control short and long term agonist-promoted desensitization of alpha 2C10, alpha 2C4, and alpha 2C2. J Biol Chem. 1992 Dec 15;267(35):25473–25479. [PubMed] [Google Scholar]
  6. Firsov D., Mandon B., Morel A., Merot J., Le Maout S., Bellanger A. C., de Rouffignac C., Elalouf J. M., Buhler J. M. Molecular analysis of vasopressin receptors in the rat nephron. Evidence for alternative splicing of the V2 receptor. Pflugers Arch. 1994 Nov;429(1):79–89. doi: 10.1007/BF02584033. [DOI] [PubMed] [Google Scholar]
  7. Forray C., Bard J. A., Wetzel J. M., Chiu G., Shapiro E., Tang R., Lepor H., Hartig P. R., Weinshank R. L., Branchek T. A. The alpha 1-adrenergic receptor that mediates smooth muscle contraction in human prostate has the pharmacological properties of the cloned human alpha 1c subtype. Mol Pharmacol. 1994 Apr;45(4):703–708. [PubMed] [Google Scholar]
  8. Graham R. M., Perez D. M., Hwa J., Piascik M. T. alpha 1-adrenergic receptor subtypes. Molecular structure, function, and signaling. Circ Res. 1996 May;78(5):737–749. doi: 10.1161/01.res.78.5.737. [DOI] [PubMed] [Google Scholar]
  9. Grosse R., Schöneberg T., Schultz G., Gudermann T. Inhibition of gonadotropin-releasing hormone receptor signaling by expression of a splice variant of the human receptor. Mol Endocrinol. 1997 Aug;11(9):1305–1318. doi: 10.1210/mend.11.9.9966. [DOI] [PubMed] [Google Scholar]
  10. Hebert T. E., Loisel T. P., Adam L., Ethier N., Onge S. S., Bouvier M. Functional rescue of a constitutively desensitized beta2AR through receptor dimerization. Biochem J. 1998 Feb 15;330(Pt 1):287–293. doi: 10.1042/bj3300287. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hebert T. E., Moffett S., Morello J. P., Loisel T. P., Bichet D. G., Barret C., Bouvier M. A peptide derived from a beta2-adrenergic receptor transmembrane domain inhibits both receptor dimerization and activation. J Biol Chem. 1996 Jul 5;271(27):16384–16392. doi: 10.1074/jbc.271.27.16384. [DOI] [PubMed] [Google Scholar]
  12. Hieble J. P., Bylund D. B., Clarke D. E., Eikenburg D. C., Langer S. Z., Lefkowitz R. J., Minneman K. P., Ruffolo R. R., Jr International Union of Pharmacology. X. Recommendation for nomenclature of alpha 1-adrenoceptors: consensus update. Pharmacol Rev. 1995 Jun;47(2):267–270. [PubMed] [Google Scholar]
  13. Hirasawa A., Horie K., Tanaka T., Takagaki K., Murai M., Yano J., Tsujimoto G. Cloning, functional expression and tissue distribution of human cDNA for the alpha 1C-adrenergic receptor. Biochem Biophys Res Commun. 1993 Sep 15;195(2):902–909. doi: 10.1006/bbrc.1993.2130. [DOI] [PubMed] [Google Scholar]
  14. Hirasawa A., Shibata K., Horie K., Takei Y., Obika K., Tanaka T., Muramoto N., Takagaki K., Yano J., Tsujimoto G. Cloning, functional expression and tissue distribution of human alpha 1c-adrenoceptor splice variants. FEBS Lett. 1995 Apr 24;363(3):256–260. doi: 10.1016/0014-5793(95)00330-c. [DOI] [PubMed] [Google Scholar]
  15. Hirasawa A., Sugawara T., Awaji T., Tsumaya K., Ito H., Tsujimoto G. Subtype-specific differences in subcellular localization of alpha1-adrenoceptors: chlorethylclonidine preferentially alkylates the accessible cell surface alpha1-adrenoceptors irrespective of the subtype. Mol Pharmacol. 1997 Nov;52(5):764–770. doi: 10.1124/mol.52.5.764. [DOI] [PubMed] [Google Scholar]
  16. Hirasawa A., Tsumaya K., Awaji T., Shibata K., Homma N., Shinomiya T., Tsujimoto G. Flow cytometry analysis of alpha1-adrenoceptor subtypes. FEBS Lett. 1996 May 20;386(2-3):141–148. doi: 10.1016/0014-5793(96)00388-2. [DOI] [PubMed] [Google Scholar]
  17. Kyte J., Doolittle R. F. A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982 May 5;157(1):105–132. doi: 10.1016/0022-2836(82)90515-0. [DOI] [PubMed] [Google Scholar]
  18. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  19. Liu K., Bergson C., Levenson R., Schmauss C. On the origin of mRNA encoding the truncated dopamine D3-type receptor D3nf and detection of D3nf-like immunoreactivity in human brain. J Biol Chem. 1994 Nov 18;269(46):29220–29226. [PubMed] [Google Scholar]
  20. Marshall I., Burt R. P., Chapple C. R. Noradrenaline contractions of human prostate mediated by alpha 1A-(alpha 1c-) adrenoceptor subtype. Br J Pharmacol. 1995 Jul;115(5):781–786. doi: 10.1111/j.1476-5381.1995.tb15001.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Moriyama N., Yamaguchi T., Takeuchi T., Sakamoto E., Ueki T., Tsujimoto G., Kawabe K. Semiquantitative evaluation of alpha1A-adrenoceptor subtype mRNA in human hypertrophied and non-hypertrophied prostates: regional comparison. Life Sci. 1999;64(3):201–210. doi: 10.1016/s0024-3205(98)00552-9. [DOI] [PubMed] [Google Scholar]
  22. Nakamura M., Sakanaka C., Aoki Y., Ogasawara H., Tsuji T., Kodama H., Matsumoto T., Shimizu T., Noma M. Identification of two isoforms of mouse neuropeptide Y-Y1 receptor generated by alternative splicing. Isolation, genomic structure, and functional expression of the receptors. J Biol Chem. 1995 Dec 15;270(50):30102–30110. doi: 10.1074/jbc.270.50.30102. [DOI] [PubMed] [Google Scholar]
  23. Nimchinsky E. A., Hof P. R., Janssen W. G., Morrison J. H., Schmauss C. Expression of dopamine D3 receptor dimers and tetramers in brain and in transfected cells. J Biol Chem. 1997 Nov 14;272(46):29229–29237. doi: 10.1074/jbc.272.46.29229. [DOI] [PubMed] [Google Scholar]
  24. Ramarao C. S., Denker J. M., Perez D. M., Gaivin R. J., Riek R. P., Graham R. M. Genomic organization and expression of the human alpha 1B-adrenergic receptor. J Biol Chem. 1992 Oct 25;267(30):21936–21945. [PubMed] [Google Scholar]
  25. Razik M. A., Lee K., Price R. R., Williams M. R., Ongjoco R. R., Dole M. K., Rudner X. L., Kwatra M. M., Schwinn D. A. Transcriptional regulation of the human alpha1a-adrenergic receptor gene. Characterization Of the 5'-regulatory and promoter region. J Biol Chem. 1997 Nov 7;272(45):28237–28246. doi: 10.1074/jbc.272.45.28237. [DOI] [PubMed] [Google Scholar]
  26. Rokosh D. G., Stewart A. F., Chang K. C., Bailey B. A., Karliner J. S., Camacho S. A., Long C. S., Simpson P. C. Alpha1-adrenergic receptor subtype mRNAs are differentially regulated by alpha1-adrenergic and other hypertrophic stimuli in cardiac myocytes in culture and in vivo. Repression of alpha1B and alpha1D but induction of alpha1C. J Biol Chem. 1996 Mar 8;271(10):5839–5843. doi: 10.1074/jbc.271.10.5839. [DOI] [PubMed] [Google Scholar]
  27. Schmauss C. Enhanced cleavage of an atypical intron of dopamine D3-receptor pre-mRNA in chronic schizophrenia. J Neurosci. 1996 Dec 15;16(24):7902–7909. doi: 10.1523/JNEUROSCI.16-24-07902.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Schmauss C., Haroutunian V., Davis K. L., Davidson M. Selective loss of dopamine D3-type receptor mRNA expression in parietal and motor cortices of patients with chronic schizophrenia. Proc Natl Acad Sci U S A. 1993 Oct 1;90(19):8942–8946. doi: 10.1073/pnas.90.19.8942. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Schoneberg T., Yun J., Wenkert D., Wess J. Functional rescue of mutant V2 vasopressin receptors causing nephrogenic diabetes insipidus by a co-expressed receptor polypeptide. EMBO J. 1996 Mar 15;15(6):1283–1291. [PMC free article] [PubMed] [Google Scholar]
  30. Takebe Y., Seiki M., Fujisawa J., Hoy P., Yokota K., Arai K., Yoshida M., Arai N. SR alpha promoter: an efficient and versatile mammalian cDNA expression system composed of the simian virus 40 early promoter and the R-U5 segment of human T-cell leukemia virus type 1 long terminal repeat. Mol Cell Biol. 1988 Jan;8(1):466–472. doi: 10.1128/mcb.8.1.466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Theroux T. L., Esbenshade T. A., Peavy R. D., Minneman K. P. Coupling efficiencies of human alpha 1-adrenergic receptor subtypes: titration of receptor density and responsiveness with inducible and repressible expression vectors. Mol Pharmacol. 1996 Nov;50(5):1376–1387. [PubMed] [Google Scholar]
  32. Tseng-Crank J., Kost T., Goetz A., Hazum S., Roberson K. M., Haizlip J., Godinot N., Robertson C. N., Saussy D. The alpha 1C-adrenoceptor in human prostate: cloning, functional expression, and localization to specific prostatic cell types. Br J Pharmacol. 1995 Aug;115(8):1475–1485. doi: 10.1111/j.1476-5381.1995.tb16640.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Weinberg D. H., Trivedi P., Tan C. P., Mitra S., Perkins-Barrow A., Borkowski D., Strader C. D., Bayne M. Cloning, expression and characterization of human alpha adrenergic receptors alpha 1a, alpha 1b and alpha 1c. Biochem Biophys Res Commun. 1994 Jun 30;201(3):1296–1304. doi: 10.1006/bbrc.1994.1845. [DOI] [PubMed] [Google Scholar]
  34. Zhu X., Wess J. Truncated V2 vasopressin receptors as negative regulators of wild-type V2 receptor function. Biochemistry. 1998 Nov 10;37(45):15773–15784. doi: 10.1021/bi981162z. [DOI] [PubMed] [Google Scholar]

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