Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1986 Jan;83(2):498–502. doi: 10.1073/pnas.83.2.498

An acetylcholine receptor precursor alpha subunit that binds alpha-bungarotoxin but not d-tubocurare.

B E Carlin, J C Lawrence Jr, J M Lindstrom, J P Merlie
PMCID: PMC322887  PMID: 3455784

Abstract

We have identified an intracellular form of the alpha subunit of the acetylcholine receptor that binds alpha-bungarotoxin with high affinity. Unlike the mature receptor complex, an alpha 2 beta gamma delta pentamer that migrates as a 9S species in velocity sedimentation analysis, the intracellular species moves as a 5S component. The kinetics of appearance of alpha subunit in the 5S component and the mature receptor complex indicate that the intracellular 5S component is a precursor of the mature receptor. The precursor species differs from 9S receptor in two critical features: (i) the precursor alpha subunit is not associated with beta subunit and (ii) alpha-bungarotoxin binding to the precursor alpha subunit is not inhibited by the cholinergic ligands decamethonium or d-tubocurarine. The properties of the precursor suggest that the acquisition of the ligand binding site by alpha subunit occurs at a distinct stage in the posttranslational development of functional acetylcholine receptor.

Full text

PDF
498

Images in this article

500Image
on p.500
500Image
on p.500

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson D. J., Blobel G. Molecular events in the synthesis and assembly of a nicotinic acetylcholine receptor. Cold Spring Harb Symp Quant Biol. 1983;48(Pt 1):125–134. doi: 10.1101/sqb.1983.048.01.015. [DOI] [PubMed] [Google Scholar]
  2. Boulter J., Luyten W., Evans K., Mason P., Ballivet M., Goldman D., Stengelin S., Martin G., Heinemann S., Patrick J. Isolation of a clone coding for the alpha-subunit of a mouse acetylcholine receptor. J Neurosci. 1985 Sep;5(9):2545–2552. doi: 10.1523/JNEUROSCI.05-09-02545.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Devreotes P. N., Fambrough D. M. Acetylcholine receptor turnover in membranes of developing muscle fibers. J Cell Biol. 1975 May;65(2):335–358. doi: 10.1083/jcb.65.2.335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Devreotes P. N., Gardner J. M., Fambrough D. M. Kinetics of biosynthesis of acetylcholine receptor and subsequent incorporation into plasma membrane of cultured chick skeletal muscle. Cell. 1977 Mar;10(3):365–373. doi: 10.1016/0092-8674(77)90023-x. [DOI] [PubMed] [Google Scholar]
  5. Gershoni J. M., Hawrot E., Lentz T. L. Binding of alpha-bungarotoxin to isolated alpha subunit of the acetylcholine receptor of Torpedo californica: quantitative analysis with protein blots. Proc Natl Acad Sci U S A. 1983 Aug;80(16):4973–4977. doi: 10.1073/pnas.80.16.4973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Gullick W. J., Lindstrom J. M. Mapping the binding of monoclonal antibodies to the acetylcholine receptor from Torpedo californica. Biochemistry. 1983 Jul 5;22(14):3312–3320. doi: 10.1021/bi00283a003. [DOI] [PubMed] [Google Scholar]
  7. Haggerty J. G., Froehner S. C. Restoration of 125I-alpha-bungarotoxin binding activity to the alpha subunit of Torpedo acetylcholine receptor isolated by gel electrophoresis in sodium dodecyl sulfate. J Biol Chem. 1981 Aug 25;256(16):8294–8297. [PubMed] [Google Scholar]
  8. Kao P. N., Dwork A. J., Kaldany R. R., Silver M. L., Wideman J., Stein S., Karlin A. Identification of the alpha subunit half-cystine specifically labeled by an affinity reagent for the acetylcholine receptor binding site. J Biol Chem. 1984 Oct 10;259(19):11662–11665. [PubMed] [Google Scholar]
  9. Lawrence J. C., Catterall W. A. Tetrodotoxin-insensitive sodium channels. Binding of polypeptide neurotoxins in primary cultures of rat muscle cells. J Biol Chem. 1981 Jun 25;256(12):6223–6229. [PubMed] [Google Scholar]
  10. Lindstrom J., Gullick W., Conti-Tronconi B., Ellisman M. Proteolytic nicking of the acetylcholine receptor. Biochemistry. 1980 Oct 14;19(21):4791–4795. doi: 10.1021/bi00562a012. [DOI] [PubMed] [Google Scholar]
  11. Lo M. M., Dolly J. O., Barnard E. A. Molecular forms of the acetylcholine receptor from vertebrate muscles and Torpedo electric organ. Interactions with specific ligands. Eur J Biochem. 1981 May;116(1):155–163. doi: 10.1111/j.1432-1033.1981.tb05313.x. [DOI] [PubMed] [Google Scholar]
  12. Merlie J. P. Biogenesis of the acetylcholine receptor, a multisubunit integral membrane protein. Cell. 1984 Mar;36(3):573–575. doi: 10.1016/0092-8674(84)90335-0. [DOI] [PubMed] [Google Scholar]
  13. Merlie J. P., Isenberg K. E., Russell S. D., Sanes J. R. Denervation supersensitivity in skeletal muscle: analysis with a cloned cDNA probe. J Cell Biol. 1984 Jul;99(1 Pt 1):332–335. doi: 10.1083/jcb.99.1.332. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Merlie J. P., Lindstrom J. Assembly in vivo of mouse muscle acetylcholine receptor: identification of an alpha subunit species that may be an assembly intermediate. Cell. 1983 Oct;34(3):747–757. doi: 10.1016/0092-8674(83)90531-7. [DOI] [PubMed] [Google Scholar]
  15. Merlie J. P., Sebbane R. Acetylcholine receptor subunits transit a precursor pool before acquiring alpha-bungarotoxin binding activity. J Biol Chem. 1981 Apr 25;256(8):3605–3608. [PubMed] [Google Scholar]
  16. Merlie J. P., Sebbane R., Gardner S., Lindstrom J. cDNA clone for the alpha subunit of the acetylcholine receptor from the mouse muscle cell line BC3H-1. Proc Natl Acad Sci U S A. 1983 Jun;80(12):3845–3849. doi: 10.1073/pnas.80.12.3845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Merlie J. P., Sebbane R., Gardner S., Olson E., Lindstrom J. The regulation of acetylcholine receptor expression in mammalian muscle. Cold Spring Harb Symp Quant Biol. 1983;48(Pt 1):135–146. doi: 10.1101/sqb.1983.048.01.016. [DOI] [PubMed] [Google Scholar]
  18. Merlie J. P., Sebbane R., Tzartos S., Lindstrom J. Inhibition of glycosylation with tunicamycin blocks assembly of newly synthesized acetylcholine receptor subunits in muscle cells. J Biol Chem. 1982 Mar 10;257(5):2694–2701. [PubMed] [Google Scholar]
  19. Mishina M., Tobimatsu T., Imoto K., Tanaka K., Fujita Y., Fukuda K., Kurasaki M., Takahashi H., Morimoto Y., Hirose T. Location of functional regions of acetylcholine receptor alpha-subunit by site-directed mutagenesis. 1985 Jan 31-Feb 6Nature. 313(6001):364–369. doi: 10.1038/313364a0. [DOI] [PubMed] [Google Scholar]
  20. Oblas B., Boyd N. D., Singer R. H. Analysis of receptor-ligand interactions using nitrocellulose gel transfer: application to Torpedo acetylcholine receptor and alpha-bungarotoxin. Anal Biochem. 1983 Apr 1;130(1):1–8. doi: 10.1016/0003-2697(83)90641-3. [DOI] [PubMed] [Google Scholar]
  21. Peper K., Bradley R. J., Dreyer F. The acetylcholine receptor at the neuromuscular junction. Physiol Rev. 1982 Oct;62(4 Pt 1):1271–1340. doi: 10.1152/physrev.1982.62.4.1271. [DOI] [PubMed] [Google Scholar]
  22. Popot J. L., Changeux J. P. Nicotinic receptor of acetylcholine: structure of an oligomeric integral membrane protein. Physiol Rev. 1984 Oct;64(4):1162–1239. doi: 10.1152/physrev.1984.64.4.1162. [DOI] [PubMed] [Google Scholar]
  23. Shorr R. G., Lyddiatt A., Lo M. M., Dolly J. O., Barnard E. A. Acetylcholine receptor from mammalian skeletal muscle. Oligomeric forms and their subunit structures. Eur J Biochem. 1981 May;116(1):143–153. doi: 10.1111/j.1432-1033.1981.tb05312.x. [DOI] [PubMed] [Google Scholar]
  24. Tzartos S. J., Changeux J. P. High affinity binding of alpha-bungarotoxin to the purified alpha-subunit and to its 27,000-dalton proteolytic peptide from Torpedo marmorata acetylcholine receptor. Requirement for sodium dodecyl sulfate. EMBO J. 1983;2(3):381–387. doi: 10.1002/j.1460-2075.1983.tb01434.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tzartos S. J., Rand D. E., Einarson B. L., Lindstrom J. M. Mapping of surface structures of electrophorus acetylcholine receptor using monoclonal antibodies. J Biol Chem. 1981 Aug 25;256(16):8635–8645. [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES