Two distinct gating mechanisms in gap junction channels: CO2-sensitive and voltage-sensitive

Bukauskas, Feliksas F.; Peracchia, Camillo

doi:10.1016/s0006-3495(97)78856-8

Cited by 111 publications

(112 citation statements)

References 24 publications

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“…Analogous to the TSF system of Harris et al (37) or the excised patch experiments of Verselis et al (38), this is a reductionist system. We limit the number of variables in the imaging experiment, which makes our results complementary to the pH studies done in whole cells (12,39). We have found that Cx26 hemichannels close in response to acidification, but only in the presence of an aminosulfonate buffer, suggesting that the aminosulfonate cation acts as a modulator or ligand to specific sites on the Cx26 protein.…”

Section: Discussionmentioning

confidence: 66%

Aminosulfonate Modulated pH-induced Conformational Changes in Connexin26 Hemichannels

Bippes

Hand

et al. 2007

Journal of Biological Chemistry

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Gap junction channels regulate cell-cell communication by passing metabolites, ions, and signaling molecules. Gap junction channel closure in cells by acidification is well documented; however, it is unknown whether acidification affects connexins or modulating proteins or compounds that in turn act on connexins. Protonated aminosulfonates directly inhibit connexin channel activity in an isoform-specific manner as shown in previously published studies. High-resolution atomic force microscopy of force-dissected connexin26 gap junctions revealed that in HEPES buffer, the pore was closed at pH < 6.5 and opened reversibly by increasing the pH to 7.6. This pH effect was not observed in non-aminosulfonate buffers. Increasing the protonated HEPES concentration did not close the pore, indicating that a saturation of the binding sites occurs at 10 mM HEPES. Analysis of the extracellular surface topographs reveals that the pore diameter increases gradually with pH. The outer connexon diameter remains unchanged, and there is a ϳ6.5°rotation in connexon lobes. These observations suggest that the underlying mechanism closing the pore is different from an observed Ca 2؉ -induced closure.Gap junction channels (GJC) 3 are dynamic macromolecular complexes capable of opening and closing the channel pore in response to a number of stimuli such as divalent cations, signaling molecules, phosphorylation, pH, and modulators of specific isoforms (1). These regulated conduits for the passage of small molecules greatly influence homeostasis, development, ionic transmission, and other cellular processes. Whereas there exist strong cell biological, biochemical, and biophysical evidence for the effects of these modulators, there is not much information at the structural level as to the conformational changes that occur in closing the pore in response to these stimuli.Each connexin (Cx) channel is composed of two hexamers (connexons) that dock at their apposed extracellular surfaces. The cyclic arrangement of the subunits within the hexamers suggests that gating can occur by a rotation and translation of the transmembrane segments within all six monomers. It has been postulated that gating occurs as a "camera iris" shutter (2). An alternate hypothesis has been proposed in which intra-connexin associations occur to produce either a particle-receptor blockage at the cytoplasmic surface (3, 4) or as a physical gate near the extracellular surface ("loop gate") (5). Whether these proposed mechanisms correlate to the closure of fast and/or slow gates that have been characterized by electrophysiological methods (see Ref. 6) remain to be determined.Gating by intracellular acidification is one way that connexin channels open and close in response to stimuli. Experimentally determined decreases in intracellular pH are known to decrease junctional electrical coupling in cardiomyocytes and in Purkinje fibers (7-10) as well as in teleost and amphibian embryos (11). Stergiopoulos et al. (12) showed that many, but not all, connexins close in a pH-sensit...

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Section: Discussionmentioning

confidence: 66%

Aminosulfonate Modulated pH-induced Conformational Changes in Connexin26 Hemichannels

Bippes

Hand

et al. 2007

Journal of Biological Chemistry

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“…At least two voltage sensitive gates have been identified: fast and slow. The fast gate and chemical gate are believed to be distinct, whereas slow gate and chemical gate are likely to be the same (Bukauskas and Peracchia, 1997;Peracchia et al, 1999Peracchia et al, , 2000. Slow and fast V j gates are in series and each hemichannel appears to have both gates.…”

Section: B Effects Of Cftr Expression On Cx45 Voltage-sensitivity Anmentioning

confidence: 99%

Interactions of connexins with other membrane channels and transporters

Chanson

Kotsias

Peracchia

et al. 2007

Progress in Biophysics and Molecular Biology

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Cell-to-cell communication through gap junctions exists in most animal cells and is essential for many important biological processes including rapid transmission of electric signals to coordinate contraction of cardiac and smooth muscle, the intercellular propagation of Ca 2+ waves and synchronization of physiological processes between adjacent cells within a tissue. Recent studies have shown that connexins can have either direct or indirect interactions with other plasma membrane ion channels or membrane transport proteins with important functional consequences. For example, in tissues most severely affected by cystic fibrosis, activation of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) has been shown to influence connexin function. Moreover, a direct interaction between Cx45.6 and the Major Intrinsic Protein/AQP0 in lens appears to influence the process of cell differentiation whereas interactions between aquaporin 4 (AQP4) and Cx43 in mouse astrocytes may coordinate the intercellular movement of ions and water between astrocytes. In this review, we discuss evidence supporting interactions between connexins and membrane channels/ transporters including CFTR, aquaporins, ionotropic glutamate receptors, and between pannexin1, another class of putative gap-junction-forming proteins, and Kvβ3, a regulatory β-subunit of voltage gated potassium channels. Although the precise molecular nature of these interactions has yet to be defined, their consequences may be critical for normal tissue homeostasis.

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“…It has been shown that gating of GJ channels by voltage is predominantly between the open state and a long-lived substate; the V j gating transitions are fast and typical of ion channel gating and complete closures are infrequent (11,12). However, complete closures can be induced by exposure to chemical uncouplers, such as H + or alkanols (13). The transitions associated with chemical gating are slow, taking tens of milliseconds to complete, are common to cell-cell channels formed of all connexins and resemble the gating transitions induced by low pH in Cx46 hemichannels.…”

Section: Acidification-induced Transitions Varied Frommentioning

confidence: 99%

Connexin hemichannels and cell-cell channels: comparison of properties

Verselis

Trexler

Bukauskas

2000

Braz J Med Biol Res

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Connexin46 (Cx46) forms functional hemichannels in the absence of contact by an apposed hemichannel and we have used these hemichannels to study gating and permeation at the single channel level with high time resolution. Using both cell-attached and -excised patch configurations, we find that single Cx46 hemichannels exhibit some properties expected of half of a gap junction channel, as well as novel properties. Cx46 hemichannels have a large unitary conductance (~300 pS) and a relatively large pore as inferred from permeability to TEA. Both monovalent cations and anions can permeate, but cations are substantially more permeable. The open channel conductance shows marked inward rectification in symmetric salts. We find that the conductance and permeability properties of Cx46 cell-cell channels can be explained by the series addition of two hemichannels. These data suggest that the pore structures of unapposed hemichannels and cell-cell channels are conserved. Also like cell-cell channels, unapposed Cx46 hemichannels are closed by elevated levels of H + or Ca 2+ ions on the cytoplasmic face. Closure occurs in excised patches indicating that the actions of these agents do not require a soluble cytoplasmic factor. Fast (<0.5 ms) application of H + to either side of the open hemichannel causes an immediate small reduction in unitary conductance followed by complete closure with latencies that are dependent on H + concentration and side of application; sensitivity is much greater to H + on the cytoplasmic side. Closure by cytoplasmic H + does not require that the hemichannel be open. Thus, H + ions readily permeate Cx46 hemichannels, but at high enough concentration close them by acting at a cytoplasmic site(s) that causes a conformational change resulting in complete closure. Extracellular H + may permeate to act on the cytoplasmic site or act on a lower affinity extracellular site. Thus, the unapposed hemichannel is a valuable tool in addressing fundamental questions concerning the operation of gap junction channels that are difficult to answer by existing methods. The ability of Cx46, and perhaps other connexins, to form functional unapposed hemichannels that are opened by moderate depolarization may represent an unexplored role of connexins as mediators of transport across the plasma membrane.

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Two distinct gating mechanisms in gap junction channels: CO2-sensitive and voltage-sensitive

Cited by 111 publications

References 24 publications

Aminosulfonate Modulated pH-induced Conformational Changes in Connexin26 Hemichannels

Aminosulfonate Modulated pH-induced Conformational Changes in Connexin26 Hemichannels

Interactions of connexins with other membrane channels and transporters

Connexin hemichannels and cell-cell channels: comparison of properties

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