Browsing by Author "Canan, Jonathan"
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Item Cx46 hemichannel modulation by nitric oxide: Role of the fourth transmembrane helix cysteine and its possible involvement in cataract formation.(2019) Retamal, Mauricio; Orellana, Viviana; Arévalo, Nicolás; Rojas, Cristóbal; Arjona, Rodolfo; Alcaíno, Constanza; González, Wendy; Canan, Jonathan; Moraga-Amaro, Rodrigo; Stehberg, Jimmy; Reuss, Luis; Altenberg, GuillermoUnder normal conditions, connexin (Cx) hemichannels have a low open probability, which can increase under pathological conditions. Since hemichannels are permeable to relatively large molecules, their exacerbated activity has been linked to cell damage. Cx46 is highly expressed in the lens and its mutations have been associated to cataract formation, but it is unknown whether Cx46 has a role in non-genetic cataract formation (i.e. aging and diabetes). Nitric oxide (NO) is a key element in non-genetic cataract formation and Cx46 hemichannels have been shown to be sensitive to NO. The molecular mechanisms of the effects of NO on Cx46 are unknown, but are likely to result from Cx46 S-nitrosation (also known as S-nitrosylation). In this work, we found that lens opacity was correlated with Cx46 S-nitrosation in an animal model of cataract. Consistent with this result, a NO donor increased Cx46 S-nitrosation and hemichannel opening in HLE-B3 cells (cell line derived from human lens epithelial cells). Mutagenesis studies point to the cysteine located in the fourth transmembrane helix (TM4; human C212, rat C218) as the NO sensor. Electrophysiological studies performed in Xenopus oocytes revealed that rat Cx46 hemichannels are sensitive to different NO donors, and that the presence of C218 is necessary to observe the NO donors' effects. Unexpectedly, gap junctions formed by Cx46 were insensitive to NO or the reducing agent dithiothreitol. We propose that increased hemichannel opening and/or changes in their electrophysiological properties of human Cx46 due to S-nitrosation of the cysteine in TM4 could be an important factor in cataract formation.Item Gap-junctional channel and hemichannel activity of two recently identified connexin 26 mutants associated with deafness(Springer, 2016) Dalamon, Viviana; Fiori, Mariana; Figueroa, Vania; Oliva, Carolina; Del Río, Rodrigo; González, Wendy; Canan, Jonathan; Elgoyhen, Ana; Altenberg, Guillermo; Retamal, MauricioGap-junction channels (GJCs) are formed by head-to-head association of two hemichannels (HCs, connexin hexamers). HCs and GJCs are permeable to ions and hydrophilic molecules of up to Mr ~1 kDa. Hearing impairment of genetic origin is common, and mutations of connexin 26 (Cx26) are its major cause. We recently identified two novel Cx26 mutations in hearing-impaired subjects, L10P and G109V. L10P forms functional GJCs with slightly altered voltage dependence and HCs with decrease ATP/cationic dye selectivity. G109V does not form functional GJCs, but forms functional HCs with enhanced extracellular Ca2+ sensitivity and subtle alterations in voltage dependence and ATP/cationic dye selectivity. Deafness associated with G109V could result from decreased GJCs activity, whereas deafness associated to L10P may have a more complex mechanism that involves changes in HC permeability.