HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 279, Issue 11, 9689-9692, March 12, 2004

This Article Full Text Full Text (PDF) All Versions of this Article: 279/11/9689    most recent M311438200v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Bao, X. Articles by Altenberg, G. A. Search for Related Content PubMed PubMed Citation Articles by Bao, X. Articles by Altenberg, G. A. Social Bookmarking                      What's this?

Functional Expression in Xenopus Oocytes of Gap-junctional Hemichannels Formed by a Cysteine-less Connexin 43^*

Xiaoyong Bao, Yongyue Chen, Luis Reuss, and Guillermo A. Altenberg

From the Membrane Protein Laboratory, Sealy Center for Structural Biology, and Department of Physiology and Biophysics, The University of Texas Medical Branch, Galveston, Texas 77555-0437

Gap-junctional channels are formed by two connexons or gap-junctional hemichannels in series, with each connexon conformed by six connexin molecules. As with other membrane proteins, structural information on connexons can potentially be obtained with techniques that take advantage of the highly specific thiol chemistry by positioning Cys residues at locations of interest, ideally in an otherwise Cys-less protein. It has been shown that conserved Cys residues located in the extracellular loops of connexins are essential for the docking of connexons from adjacent cells, preventing the formation of functional gap-junctional channels. Here we engineered a Cys-less version of connexin 43 (Cx43) and assessed its function using a Xenopus oocyte expression system. The Cys-less protein was expressed at the plasma membrane and did not form gap-junctional channels but formed hemichannels that behave similarly to those formed by Cx43 in terms of permeation to carboxyfluorescein. The carboxyfluorescein permeability of Cys-less hemichannels was increased by protein kinase C inhibition, like the wild-type Cx43 hemichannels. We generated a protein with a single Cys in a position (residue 34) thought to face the channel pore and show that thiol modification of the Cys abolishes the carboxyfluorescein permeability. We conclude that Cysless Cx43 forms regulated functional hemichannels, and therefore Cys-less Cx43 is a useful tool for future structural studies.

Received for publication, October 17, 2003 , and in revised form, December 9, 2003.

^* This work was supported in part by American Heart Association Grant-in-aid 0050353N and a Jeane B. Kempner Scholar Award (to X. B.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Jeane B. Kempner postdoctoral fellow.

To whom correspondence and requests for reprints should be addressed. Tel.: 409-772-3026; Fax: 409-772-1301; E-mail: galtenbe{at}utmb.edu.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				D. Tong, T. Y. Li, K. E. Naus, D. Bai, and G. M. Kidder In vivo analysis of undocked connexin43 gap junction hemichannels in ovarian granulosa cells J. Cell Sci., November 15, 2007; 120(22): 4016 - 4024. [Abstract] [Full Text] [PDF]

				M. A. Retamal, K. A. Schalper, K. F. Shoji, M. V. L. Bennett, and J. C. Saez Opening of connexin 43 hemichannels is increased by lowering intracellular redox potential PNAS, May 15, 2007; 104(20): 8322 - 8327. [Abstract] [Full Text] [PDF]

				A. Lai, D.-N. Le, W. A. Paznekas, W. D. Gifford, E. W. Jabs, and A. C. Charles Oculodentodigital dysplasia connexin43 mutations result in non-functional connexin hemichannels and gap junctions in C6 glioma cells J. Cell Sci., February 1, 2006; 119(3): 532 - 541. [Abstract] [Full Text] [PDF]

				X. Bao, Y. Chen, S. H. Lee, S. C. Lee, L. Reuss, and G. A. Altenberg Membrane Transport Proteins with Complete Replacement of Transmembrane Helices with Polyalanine Sequences Remain Functional J. Biol. Chem., March 11, 2005; 280(10): 8647 - 8650. [Abstract] [Full Text] [PDF]

				D. Locke, I. V. Koreen, J. Y. Liu, and A. L. Harris Reversible Pore Block of Connexin Channels by Cyclodextrins J. Biol. Chem., May 28, 2004; 279(22): 22883 - 22892. [Abstract] [Full Text] [PDF]