Isoform Composition of Connexin Channels Determines Selectivity among Second Messengers and Uncharged Molecules*
- From the ‡Thomas C. Jenkins Department of Biophysics, Johns Hopkins University, Baltimore, Maryland 21218, the§Department of Enzyme Technology, Gesellschaft für Biotechnologische Forschung, Mascheroder Weg 1, 38124 Braunschweig, Germany, and the ‖Department of Biochemistry, Yeungnam University, 214-1 Daedong, Kyoungsan, Republic of Korea
Abstract
Intercellular connexin channels (gap junction channels) have long been thought to mediate molecular signaling between cells, but the nature of the signaling has been unclear. This study shows that connexin channels from native tissue have selective permeabilities, partially based on pore diameter, that discriminate among cytoplasmic second messenger molecules. Permeability was assessed by measurement of selective loss/retention of tracers from liposomes containing reconstituted connexin channels. The tracers employed were tritiated cyclic nucleotides and a series of oligomaltosaccharides derivatized with a small uncharged fluorescent moiety. The data define different size cut-off limits for permeability through homomeric connexin-32 channels and through heteromeric connexin-32/connexin-26 channels. Connexin-26 contributes to a narrowed pore. Both cAMP and cGMP were permeable through the homomeric connexin-32 channels. cAMP was permeable through only a fraction of the heteromeric channels. Surprisingly, cGMP was permeable through a substantially greater fraction of the heteromeric channels than was cAMP. The data suggest that isoform stoichiometry and/or arrangement within a connexin channel determines whether cyclic nucleotides can permeate, and which ones. This is the first evidence for connexin-specific selectivity among biological signaling molecules.
Footnotes
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↵* This work was supported in part by National Institutes of Health Grant GM36044 and Office of Naval Research Grant N00014-90-J-1960 (to A. L. H.)The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵¶ Present address: Bundesministerium für Bildung, Wissenschaft, Forschung und Technologie, 53175 Bonn, Germany.
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↵** To whom correspondence should be addressed.
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↵1 The abbreviations used are: Cx, connexin; α(1→4)Glcn, α(1→4)-linked glucose linear oligomer, where n is the number of glucose units; DSP, dithiobis(succinimidyl propionate); DTSSP, dithiobis(sulfosuccinimidyl propionate); DTT, dithiothreitol; HPLC, high performance liquid chromatography; mouseCx, connexin immunopurified from mouse liver (Cx32/Cx26); PA; aminopyridyl moiety; PA-sugar; oligomaltose labeled with an aminopyridyl group; n-PA, PA-sugar wheren is the number of saccharide units; PBS, phosphate-buffered saline; ratCx, connexin immunopurified from rat liver (Cx32); TSF, transport-specific fractionation of liposomes.
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- Received June 25, 1997.
- Revision received November 3, 1997.
- The American Society for Biochemistry and Molecular Biology, Inc.
