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The opening of connexin (Cx) hemichannels in the membrane is tightly regulated by calcium (Ca2+) and membrane voltage. Electrophysiological and atomic force microscopy experiments indicate that Ca2+ stabilizes the hemichannel closed state. However, structural data show that Ca2+ binding induces an electrostatic seal preventing ion transport without significant structural rearrangements. In agreement with the closed-state stabilization hypothesis, we found that the apparent Ca2+ sensitivity is increased as the voltage is made more negative. Moreover, the voltage and Ca2+ dependence of the channel kinetics indicate that the voltage sensor movement and Ca2+ binding are allosterically coupled. An allosteric kinetic model in which the Ca2+ decreases the energy necessary to deactivate the voltage sensor reproduces the effects of Ca2+ and voltage in Cx46 hemichannels. In agreement with the model and suggesting a conformational change that narrows the pore, Ca2+ inhibits the water flux through Cx hemichannels. We conclude that Ca2+ and voltage act allosterically to stabilize the closed conformation of Cx46 hemichannels.

Indexación: Scopus. Connexin-based channels comprise hemichannels and gap junction channels. The opening of hemichannels allow for the flux of ions and molecules from the extracellular space into the cell and vice versa. Similarly, the opening of gap junction channels permits the diffusional exchange of ions and molecules between the cytoplasm and contacting cells. The controlled opening of hemichannels has been associated with several physiological cellular processes; thereby unregulated hemichannel activity may induce loss of cellular homeostasis and cell death. Hemichannel activity can be regulated through several mechanisms, such as phosphorylation, divalent cations and changes in membrane potential. Additionally, it was recently postulated that redox molecules could modify hemichannels properties in vitro. However, the molecular mechanism by which redox molecules interact with hemichannels is poorly understood. In this work, we discuss the current knowledge on connexin redox regulation and we propose the hypothesis that extracellular cysteines could be important for sensing changes in redox potential. Future studies on this topic will offer new insight into hemichannel function, thereby expanding the understanding of the contribution of hemichannels to disease progression. http://journal.frontiersin.org/article/10.3389/fphys.2016.00001/full

Connexins (Cxs) are transmembrane proteins involved in the electrical coupling of cells, the release of signaling molecules, and cell proliferation among others. These proteins are expressed in almost every tissue of the human body and mutations in these proteins are related to several hereditary diseases. Cxs monomers oligomerize in hexamers, which traffic to the plasma membrane and form connexons or hemichannels that can act as plasma membrane channels or can travel to intercellular contact zones where they form intercellular channels known as gap junctions. The opening of the Cx hemichannels in the membrane is tightly regulated by Ca2+ and membrane voltage. This regulation prevents leakage of cellular content and its malfunction may lead to pathologic conditions. In this work we studied the reciprocal regulation of Cxs hemichannels by Ca2+ and voltage. We expressed Cx46 in Xenopus laevis oocytes, and using two electrode voltage clamp, analyzed the inhibition of currents by Ca2+. We observed that Cxs inhibition by Ca2+ is voltage dependent. The apparent Ca2+ sensitivity is increased as the membrane voltage is made more negative. This change in the affinity can be explained by a stabilization of the closed state of the Cx channel by Ca2+. We propose a model in which the calcium-bonded state prevents the opening of the hemichannel. The model allows us to determine the affinity of the closed state for calcium. In agreement with the model and suggesting a stabilization of a conformational change that narrows the pore, calcium inhibits the water flux trough Cx hemichannels. This work supports the idea that Ca2+ and voltage act synergistically to promote the closing of the pore in Cx channels.