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Molecular Pharmacology

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Research ArticleArticle

β3 Subunits Promote Expression and Nicotine-Induced Up-Regulation of Human Nicotinic α6* Nicotinic Acetylcholine Receptors Expressed in Transfected Cell Lines

Prem Tumkosit, Alexander Kuryatov, Jie Luo and Jon Lindstrom
Molecular Pharmacology October 2006, 70 (4) 1358-1368; DOI: https://doi.org/10.1124/mol.106.027326
Prem Tumkosit
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Alexander Kuryatov
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Jie Luo
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Jon Lindstrom
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Abstract

Nicotinic acetylcholine receptors (AChRs) containing α6 subunits are typically found at aminergic nerve endings where they play important roles in nicotine addiction and Parkinson's disease. α6* AChRs usually contain β3 subunits. β3 subunits are presumed to assemble only in the accessory subunit position within AChRs where they do not participate in forming acetylcholine binding sites. Assembly of subunits in the accessory position may be a critical final step in assembly of mature AChRs. Human α6 AChRs subtypes were permanently transfected into human tsA201 human embryonic kidney (HEK) cell lines. α6β2β3 and α6β4β3 cell lines were found to express much larger amounts of AChRs and were more sensitive to nicotine-induced increase in the amount of AChRs than were α6β2 or α6β4 cell lines. The increased sensitivity to nicotine-induced up-regulation was due not to a β3-induced increase in affinity for nicotine but probably to a direct effect on assembly of AChR subunits. HEK cells express only a small amount of mature α6β2 AChRs, but many of these subunits are on the cell surface. This contrasts with Xenopus laevis oocytes, which express a large amount of incorrectly assembled α6β2 subunits that bind cholinergic ligands but form large amorphous intracellular aggregates. Monoclonal antibodies (mAbs) were made to the α6 and β3 subunits to aid in the characterization of these AChRs. The α6 mAbs bind to epitopes C-terminal of the extracellular domain. These data demonstrate that both cell type and the accessory subunit β3 can play important roles in α6* AChR expression, stability, and up-regulation by nicotine.

Footnotes

  • This work was supported by grant NS11323 from the National Institutes of Health and the Phillip Morris External Research Program (to J.L.).

  • P.M., A.K., and J.L. contributed equally to this work.

  • ABBREVIATIONS: AChR, acetylcholine receptor; ACh, acetylcholine; HEK, human embryonic kidney; DMEM, Dulbecco's modified Eagle's medium; PBS, phosphate-buffered saline; mAb, monoclonal antibody; TPBS, 0.5% Triton X-100 in phosphate-buffered saline; MIR, main immunogenic region.

    • Received June 1, 2006.
    • Accepted July 11, 2006.
  • The American Society for Pharmacology and Experimental Therapeutics
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Molecular Pharmacology: 70 (4)
Molecular Pharmacology
Vol. 70, Issue 4
1 Oct 2006
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Research ArticleArticle

β3 Subunits Promote Expression and Nicotine-Induced Up-Regulation of Human Nicotinic α6* Nicotinic Acetylcholine Receptors Expressed in Transfected Cell Lines

Prem Tumkosit, Alexander Kuryatov, Jie Luo and Jon Lindstrom
Molecular Pharmacology October 1, 2006, 70 (4) 1358-1368; DOI: https://doi.org/10.1124/mol.106.027326

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Research ArticleArticle

β3 Subunits Promote Expression and Nicotine-Induced Up-Regulation of Human Nicotinic α6* Nicotinic Acetylcholine Receptors Expressed in Transfected Cell Lines

Prem Tumkosit, Alexander Kuryatov, Jie Luo and Jon Lindstrom
Molecular Pharmacology October 1, 2006, 70 (4) 1358-1368; DOI: https://doi.org/10.1124/mol.106.027326
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