On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells

Anibal A. Vargas, Bruno A. Cisterna, Fujiko Saavedra-Leiva, Carolina Urrutia, Luis A. Cea, Alex H. Vielma, Sebastian E. Gutierrez-Maldonado, Alberto J M Martin, Claudia Pareja-Barrueto, Yerko Escalona, Oliver Schmachtenberg, Carlos F. Lagos, Tomas Perez-Acle, Juan C. Sáez

Resultado de la investigación: Article

6 Citas (Scopus)

Resumen

Although connexins (Cxs) are broadly expressed by cells of mammalian organisms, Cx39 has a very restricted pattern of expression and the biophysical properties of Cx39-based channels [hemichannels (HCs) and gap junction channels (GJCs)] remain largely unknown. Here, we used HeLa cells transfected with Cx39 (HeLa-Cx39 cells) in which intercellular electrical coupling was not detected, indicating the absence of GJCs. However, functional HCs were found on the surface of cells exposed to conditions known to increase the open probability of other Cx HCs (e.g., extracellular divalent cationic-free solution (DCFS), extracellular alkaline pH, mechanical stimulus and depolarization to positive membrane potentials). Cx39 HCs were blocked by some traditional Cx HC blockers, but not by others or a pannexin1 channel blocker. HeLa-Cx39 cells showed similar resting membrane potentials (RMPs) to those of parental cells, and exposure to DCFS reduced RMPs in Cx39 transfectants, but not in parental cells. Under these conditions, unitary events of ~75 pS were frequent in HeLa-Cx39 cells and absent in parental cells. Real-time cellular uptake experiments of dyes with different physicochemical features, as well as the application of a machine-learning approach revealed that Cx39 HCs are preferentially permeable to molecules characterized by six categories of descriptors, namely: (1) electronegativity, (2) ionization potential, (3) polarizability, (4) size and geometry, (5) topological flexibility and (6) valence. However, Cx39 HCs opened by mechanical stimulation or alkaline pH were impermeable to Ca2+. Molecular modeling of Cx39-based channels suggest that a constriction present at the intracellular portion of the para helix region co-localizes with an electronegative patch, imposing an energetic and steric barrier, which in the case of GJCs may hinder channel function. Results reported here demonstrate that Cx39 form HCs and add to our understanding of the functional roles of Cx39 HCs under physiological and pathological conditions in cells that express them.

Idioma originalEnglish
Número de artículo38
PublicaciónFrontiers in Physiology
Volumen8
N.ºFEB
DOI
EstadoPublished - 9 feb 2017

Huella dactilar

Connexins
Gap Junctions
HeLa Cells
Membrane Potentials
Constriction
Coloring Agents

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

Citar esto

Vargas, A. A., Cisterna, B. A., Saavedra-Leiva, F., Urrutia, C., Cea, L. A., Vielma, A. H., ... Sáez, J. C. (2017). On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells. Frontiers in Physiology, 8(FEB), [38]. https://doi.org/10.3389/fphys.2017.00038
Vargas, Anibal A. ; Cisterna, Bruno A. ; Saavedra-Leiva, Fujiko ; Urrutia, Carolina ; Cea, Luis A. ; Vielma, Alex H. ; Gutierrez-Maldonado, Sebastian E. ; Martin, Alberto J M ; Pareja-Barrueto, Claudia ; Escalona, Yerko ; Schmachtenberg, Oliver ; Lagos, Carlos F. ; Perez-Acle, Tomas ; Sáez, Juan C. / On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells. En: Frontiers in Physiology. 2017 ; Vol. 8, N.º FEB.
@article{ba128926268f493a800e48591fd02c69,
title = "On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells",
abstract = "Although connexins (Cxs) are broadly expressed by cells of mammalian organisms, Cx39 has a very restricted pattern of expression and the biophysical properties of Cx39-based channels [hemichannels (HCs) and gap junction channels (GJCs)] remain largely unknown. Here, we used HeLa cells transfected with Cx39 (HeLa-Cx39 cells) in which intercellular electrical coupling was not detected, indicating the absence of GJCs. However, functional HCs were found on the surface of cells exposed to conditions known to increase the open probability of other Cx HCs (e.g., extracellular divalent cationic-free solution (DCFS), extracellular alkaline pH, mechanical stimulus and depolarization to positive membrane potentials). Cx39 HCs were blocked by some traditional Cx HC blockers, but not by others or a pannexin1 channel blocker. HeLa-Cx39 cells showed similar resting membrane potentials (RMPs) to those of parental cells, and exposure to DCFS reduced RMPs in Cx39 transfectants, but not in parental cells. Under these conditions, unitary events of ~75 pS were frequent in HeLa-Cx39 cells and absent in parental cells. Real-time cellular uptake experiments of dyes with different physicochemical features, as well as the application of a machine-learning approach revealed that Cx39 HCs are preferentially permeable to molecules characterized by six categories of descriptors, namely: (1) electronegativity, (2) ionization potential, (3) polarizability, (4) size and geometry, (5) topological flexibility and (6) valence. However, Cx39 HCs opened by mechanical stimulation or alkaline pH were impermeable to Ca2+. Molecular modeling of Cx39-based channels suggest that a constriction present at the intracellular portion of the para helix region co-localizes with an electronegative patch, imposing an energetic and steric barrier, which in the case of GJCs may hinder channel function. Results reported here demonstrate that Cx39 form HCs and add to our understanding of the functional roles of Cx39 HCs under physiological and pathological conditions in cells that express them.",
keywords = "Cx39, Dye-uptake, Electrical coupling, Gap junction, Membrane potential, Permeability, Unitary conductance",
author = "Vargas, {Anibal A.} and Cisterna, {Bruno A.} and Fujiko Saavedra-Leiva and Carolina Urrutia and Cea, {Luis A.} and Vielma, {Alex H.} and Gutierrez-Maldonado, {Sebastian E.} and Martin, {Alberto J M} and Claudia Pareja-Barrueto and Yerko Escalona and Oliver Schmachtenberg and Lagos, {Carlos F.} and Tomas Perez-Acle and S{\'a}ez, {Juan C.}",
year = "2017",
month = "2",
day = "9",
doi = "10.3389/fphys.2017.00038",
language = "English",
volume = "8",
journal = "Frontiers in Physiology",
issn = "1664-042X",
publisher = "Frontiers Media S.A.",
number = "FEB",

}

Vargas, AA, Cisterna, BA, Saavedra-Leiva, F, Urrutia, C, Cea, LA, Vielma, AH, Gutierrez-Maldonado, SE, Martin, AJM, Pareja-Barrueto, C, Escalona, Y, Schmachtenberg, O, Lagos, CF, Perez-Acle, T & Sáez, JC 2017, 'On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells', Frontiers in Physiology, vol. 8, n.º FEB, 38. https://doi.org/10.3389/fphys.2017.00038

On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells. / Vargas, Anibal A.; Cisterna, Bruno A.; Saavedra-Leiva, Fujiko; Urrutia, Carolina; Cea, Luis A.; Vielma, Alex H.; Gutierrez-Maldonado, Sebastian E.; Martin, Alberto J M; Pareja-Barrueto, Claudia; Escalona, Yerko; Schmachtenberg, Oliver; Lagos, Carlos F.; Perez-Acle, Tomas; Sáez, Juan C.

En: Frontiers in Physiology, Vol. 8, N.º FEB, 38, 09.02.2017.

Resultado de la investigación: Article

TY - JOUR

T1 - On biophysical properties and sensitivity to gap junction blockers of connexin 39 hemichannels expressed in hela cells

AU - Vargas, Anibal A.

AU - Cisterna, Bruno A.

AU - Saavedra-Leiva, Fujiko

AU - Urrutia, Carolina

AU - Cea, Luis A.

AU - Vielma, Alex H.

AU - Gutierrez-Maldonado, Sebastian E.

AU - Martin, Alberto J M

AU - Pareja-Barrueto, Claudia

AU - Escalona, Yerko

AU - Schmachtenberg, Oliver

AU - Lagos, Carlos F.

AU - Perez-Acle, Tomas

AU - Sáez, Juan C.

PY - 2017/2/9

Y1 - 2017/2/9

N2 - Although connexins (Cxs) are broadly expressed by cells of mammalian organisms, Cx39 has a very restricted pattern of expression and the biophysical properties of Cx39-based channels [hemichannels (HCs) and gap junction channels (GJCs)] remain largely unknown. Here, we used HeLa cells transfected with Cx39 (HeLa-Cx39 cells) in which intercellular electrical coupling was not detected, indicating the absence of GJCs. However, functional HCs were found on the surface of cells exposed to conditions known to increase the open probability of other Cx HCs (e.g., extracellular divalent cationic-free solution (DCFS), extracellular alkaline pH, mechanical stimulus and depolarization to positive membrane potentials). Cx39 HCs were blocked by some traditional Cx HC blockers, but not by others or a pannexin1 channel blocker. HeLa-Cx39 cells showed similar resting membrane potentials (RMPs) to those of parental cells, and exposure to DCFS reduced RMPs in Cx39 transfectants, but not in parental cells. Under these conditions, unitary events of ~75 pS were frequent in HeLa-Cx39 cells and absent in parental cells. Real-time cellular uptake experiments of dyes with different physicochemical features, as well as the application of a machine-learning approach revealed that Cx39 HCs are preferentially permeable to molecules characterized by six categories of descriptors, namely: (1) electronegativity, (2) ionization potential, (3) polarizability, (4) size and geometry, (5) topological flexibility and (6) valence. However, Cx39 HCs opened by mechanical stimulation or alkaline pH were impermeable to Ca2+. Molecular modeling of Cx39-based channels suggest that a constriction present at the intracellular portion of the para helix region co-localizes with an electronegative patch, imposing an energetic and steric barrier, which in the case of GJCs may hinder channel function. Results reported here demonstrate that Cx39 form HCs and add to our understanding of the functional roles of Cx39 HCs under physiological and pathological conditions in cells that express them.

AB - Although connexins (Cxs) are broadly expressed by cells of mammalian organisms, Cx39 has a very restricted pattern of expression and the biophysical properties of Cx39-based channels [hemichannels (HCs) and gap junction channels (GJCs)] remain largely unknown. Here, we used HeLa cells transfected with Cx39 (HeLa-Cx39 cells) in which intercellular electrical coupling was not detected, indicating the absence of GJCs. However, functional HCs were found on the surface of cells exposed to conditions known to increase the open probability of other Cx HCs (e.g., extracellular divalent cationic-free solution (DCFS), extracellular alkaline pH, mechanical stimulus and depolarization to positive membrane potentials). Cx39 HCs were blocked by some traditional Cx HC blockers, but not by others or a pannexin1 channel blocker. HeLa-Cx39 cells showed similar resting membrane potentials (RMPs) to those of parental cells, and exposure to DCFS reduced RMPs in Cx39 transfectants, but not in parental cells. Under these conditions, unitary events of ~75 pS were frequent in HeLa-Cx39 cells and absent in parental cells. Real-time cellular uptake experiments of dyes with different physicochemical features, as well as the application of a machine-learning approach revealed that Cx39 HCs are preferentially permeable to molecules characterized by six categories of descriptors, namely: (1) electronegativity, (2) ionization potential, (3) polarizability, (4) size and geometry, (5) topological flexibility and (6) valence. However, Cx39 HCs opened by mechanical stimulation or alkaline pH were impermeable to Ca2+. Molecular modeling of Cx39-based channels suggest that a constriction present at the intracellular portion of the para helix region co-localizes with an electronegative patch, imposing an energetic and steric barrier, which in the case of GJCs may hinder channel function. Results reported here demonstrate that Cx39 form HCs and add to our understanding of the functional roles of Cx39 HCs under physiological and pathological conditions in cells that express them.

KW - Cx39

KW - Dye-uptake

KW - Electrical coupling

KW - Gap junction

KW - Membrane potential

KW - Permeability

KW - Unitary conductance

UR - http://www.scopus.com/inward/record.url?scp=85014210939&partnerID=8YFLogxK

U2 - 10.3389/fphys.2017.00038

DO - 10.3389/fphys.2017.00038

M3 - Article

AN - SCOPUS:85014210939

VL - 8

JO - Frontiers in Physiology

JF - Frontiers in Physiology

SN - 1664-042X

IS - FEB

M1 - 38

ER -