The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels

E. Brady Trexler, Feliksas F. Bukauskas, Jack Kronengold, Thaddeus A. Bargiello, Vytautas Verselis

Research output: Contribution to journalArticle

105 Citations (Scopus)

Abstract

Intercellular channels formed of members of the gene family of connexins (Cxs) vary from being substantially cation selective to being anion selective. We took advantage of the ability of Cx46 to function as an unapposed hemichannel to examine the basis of Cx charge selectivity. Previously we showed Cx46 hemichannels to be large pores that predominantly conduct cations and inwardly rectify in symmetric salts, properties suggesting selectivity is influenced by fixed negative charges located toward the extracellular end of the pore. Here we demonstrate that high ionic strength solutions applied to the extracellular, but not the intracellular, side of Cx46 hemichannels substantially reduce the ratio of cation to anion permeability. Substitution of the first extracellular loop (E1) domain of Cx32, an anion-preferring Cx, reduces conductance, converts Cx46 from cation to anion preferring, and changes the I-V relation form inwardly to outwardly rectifying. These data suggest that fixed negative charges influencing selectivity in Cx46 are located in E1 and are substantially reduced and/or are replaced with positive charges from the Cx32 E1 sequence. Extending studies to Cx46 cell-cell channels, we show that they maintain a strong preference for cations, have a conductance nearly that expected by the series addition of hemichannels, but lack rectification in symmetric salts. These properties are consistent with preservation of the fixed charge region in E1 of hemichannels, which upon docking, become symmetrically placed near the center of the cell-cell channel pore. Furthermore, heterotypic cell-cell channels formed by pairing Cx46 with Cx32 or Cx43 rectify in symmetric salts in accordance with the differences in the charges we ascribed to E1. These data are consistent with charged residues in E1 facing the channel lumen and playing an important role in determining Cx channel conductance and selectivity.

Original languageEnglish (US)
Pages (from-to)3036-3051
Number of pages16
JournalBiophysical Journal
Volume79
Issue number6
StatePublished - Dec 2000

Fingerprint

Cations
Anions
Salts
Connexin 43
Connexins
Aptitude
Osmolar Concentration
Permeability
Genes
connexin 46

ASJC Scopus subject areas

  • Biophysics

Cite this

Trexler, E. B., Bukauskas, F. F., Kronengold, J., Bargiello, T. A., & Verselis, V. (2000). The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels. Biophysical Journal, 79(6), 3036-3051.

The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels. / Trexler, E. Brady; Bukauskas, Feliksas F.; Kronengold, Jack; Bargiello, Thaddeus A.; Verselis, Vytautas.

In: Biophysical Journal, Vol. 79, No. 6, 12.2000, p. 3036-3051.

Research output: Contribution to journalArticle

Trexler, EB, Bukauskas, FF, Kronengold, J, Bargiello, TA & Verselis, V 2000, 'The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels', Biophysical Journal, vol. 79, no. 6, pp. 3036-3051.
Trexler, E. Brady ; Bukauskas, Feliksas F. ; Kronengold, Jack ; Bargiello, Thaddeus A. ; Verselis, Vytautas. / The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels. In: Biophysical Journal. 2000 ; Vol. 79, No. 6. pp. 3036-3051.
@article{6db9665d034843f8a2f1dd31ec883214,
title = "The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels",
abstract = "Intercellular channels formed of members of the gene family of connexins (Cxs) vary from being substantially cation selective to being anion selective. We took advantage of the ability of Cx46 to function as an unapposed hemichannel to examine the basis of Cx charge selectivity. Previously we showed Cx46 hemichannels to be large pores that predominantly conduct cations and inwardly rectify in symmetric salts, properties suggesting selectivity is influenced by fixed negative charges located toward the extracellular end of the pore. Here we demonstrate that high ionic strength solutions applied to the extracellular, but not the intracellular, side of Cx46 hemichannels substantially reduce the ratio of cation to anion permeability. Substitution of the first extracellular loop (E1) domain of Cx32, an anion-preferring Cx, reduces conductance, converts Cx46 from cation to anion preferring, and changes the I-V relation form inwardly to outwardly rectifying. These data suggest that fixed negative charges influencing selectivity in Cx46 are located in E1 and are substantially reduced and/or are replaced with positive charges from the Cx32 E1 sequence. Extending studies to Cx46 cell-cell channels, we show that they maintain a strong preference for cations, have a conductance nearly that expected by the series addition of hemichannels, but lack rectification in symmetric salts. These properties are consistent with preservation of the fixed charge region in E1 of hemichannels, which upon docking, become symmetrically placed near the center of the cell-cell channel pore. Furthermore, heterotypic cell-cell channels formed by pairing Cx46 with Cx32 or Cx43 rectify in symmetric salts in accordance with the differences in the charges we ascribed to E1. These data are consistent with charged residues in E1 facing the channel lumen and playing an important role in determining Cx channel conductance and selectivity.",
author = "Trexler, {E. Brady} and Bukauskas, {Feliksas F.} and Jack Kronengold and Bargiello, {Thaddeus A.} and Vytautas Verselis",
year = "2000",
month = "12",
language = "English (US)",
volume = "79",
pages = "3036--3051",
journal = "Biophysical Journal",
issn = "0006-3495",
publisher = "Biophysical Society",
number = "6",

}

TY - JOUR

T1 - The first extracellular loop domain is a major determinant of charge selectivity in connexin46 channels

AU - Trexler, E. Brady

AU - Bukauskas, Feliksas F.

AU - Kronengold, Jack

AU - Bargiello, Thaddeus A.

AU - Verselis, Vytautas

PY - 2000/12

Y1 - 2000/12

N2 - Intercellular channels formed of members of the gene family of connexins (Cxs) vary from being substantially cation selective to being anion selective. We took advantage of the ability of Cx46 to function as an unapposed hemichannel to examine the basis of Cx charge selectivity. Previously we showed Cx46 hemichannels to be large pores that predominantly conduct cations and inwardly rectify in symmetric salts, properties suggesting selectivity is influenced by fixed negative charges located toward the extracellular end of the pore. Here we demonstrate that high ionic strength solutions applied to the extracellular, but not the intracellular, side of Cx46 hemichannels substantially reduce the ratio of cation to anion permeability. Substitution of the first extracellular loop (E1) domain of Cx32, an anion-preferring Cx, reduces conductance, converts Cx46 from cation to anion preferring, and changes the I-V relation form inwardly to outwardly rectifying. These data suggest that fixed negative charges influencing selectivity in Cx46 are located in E1 and are substantially reduced and/or are replaced with positive charges from the Cx32 E1 sequence. Extending studies to Cx46 cell-cell channels, we show that they maintain a strong preference for cations, have a conductance nearly that expected by the series addition of hemichannels, but lack rectification in symmetric salts. These properties are consistent with preservation of the fixed charge region in E1 of hemichannels, which upon docking, become symmetrically placed near the center of the cell-cell channel pore. Furthermore, heterotypic cell-cell channels formed by pairing Cx46 with Cx32 or Cx43 rectify in symmetric salts in accordance with the differences in the charges we ascribed to E1. These data are consistent with charged residues in E1 facing the channel lumen and playing an important role in determining Cx channel conductance and selectivity.

AB - Intercellular channels formed of members of the gene family of connexins (Cxs) vary from being substantially cation selective to being anion selective. We took advantage of the ability of Cx46 to function as an unapposed hemichannel to examine the basis of Cx charge selectivity. Previously we showed Cx46 hemichannels to be large pores that predominantly conduct cations and inwardly rectify in symmetric salts, properties suggesting selectivity is influenced by fixed negative charges located toward the extracellular end of the pore. Here we demonstrate that high ionic strength solutions applied to the extracellular, but not the intracellular, side of Cx46 hemichannels substantially reduce the ratio of cation to anion permeability. Substitution of the first extracellular loop (E1) domain of Cx32, an anion-preferring Cx, reduces conductance, converts Cx46 from cation to anion preferring, and changes the I-V relation form inwardly to outwardly rectifying. These data suggest that fixed negative charges influencing selectivity in Cx46 are located in E1 and are substantially reduced and/or are replaced with positive charges from the Cx32 E1 sequence. Extending studies to Cx46 cell-cell channels, we show that they maintain a strong preference for cations, have a conductance nearly that expected by the series addition of hemichannels, but lack rectification in symmetric salts. These properties are consistent with preservation of the fixed charge region in E1 of hemichannels, which upon docking, become symmetrically placed near the center of the cell-cell channel pore. Furthermore, heterotypic cell-cell channels formed by pairing Cx46 with Cx32 or Cx43 rectify in symmetric salts in accordance with the differences in the charges we ascribed to E1. These data are consistent with charged residues in E1 facing the channel lumen and playing an important role in determining Cx channel conductance and selectivity.

UR - http://www.scopus.com/inward/record.url?scp=0033636361&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033636361&partnerID=8YFLogxK

M3 - Article

C2 - 11106610

AN - SCOPUS:0033636361

VL - 79

SP - 3036

EP - 3051

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 6

ER -