Polydisperse molecular architecture of connexin 26/30 heteromeric hemichannels revealed by atomic force microscopy imaging

Naulin, Pamela A.; Lozano, Benjamin; Fuentes, Christian; Liu, Yu; Schmidt, Carla; Contreras, Jorge E.; Barrera, Nelson P.

doi:10.1074/jbc.ra119.012128

Cited by 5 publications

(5 citation statements)

References 44 publications

(78 reference statements)

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“…To get some molecular insights into the mechanism of action of the hyperactive HCs, molecular dynamics simulations were run for putative heteromeric HCs formed by Cx26S17F/Cx30 ( Figure 8 ). Heteromeric HCs were built for Cx26S17F/Cx30 and Cx26WT/Cx30 using the most probable stoichiometry found by Naulin et al (2020) , which the hexamer is formed by the following subunit combination: Cx26:Cx26:Cx30:Cx30:Cx26:Cx30. Both HCs, with Cx26WT and Cx26S17F, showed similar behavior, maintaining their overall structure and inner diameter.…”

Section: Resultsmentioning

confidence: 99%

“…(A,B) . Molecular dynamic simulation of a heteromeric HC composed by Cx26 or Cx26S17F and Cx30 (A) , according to the most frequent combination (stoichiometry) of each monomer in the heteromeric hemichannel (B) , as described in Naulin et al, 2020 , which is Cx26-Cx26-Cx30-Cx30-Cx26-Cx30. Colors indicate Cx26 or Cx26S17F in green and Cx30 in grey.…”

Section: Resultsmentioning

confidence: 99%

“…Missing residues and sections from original coordinates were modeled using MODELER ( Martí-Renom et al, 2000 ). The stoichiometry used was Cx26:Cx26:Cx30:Cx30:Cx26:Cx30 because it is the most abundant composition found in vitro ( Naulin et al, 2020 ). The best models were chosen using the MAIDEN program ( Postic et al, 2015 ).…”

Section: Methodsmentioning

confidence: 99%

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Expression of KID syndromic mutation Cx26S17F produces hyperactive hemichannels in supporting cells of the organ of Corti

Abbott

García²,

Villanelo³

et al. 2023

Front. Cell Dev. Biol.

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Some mutations in gap junction protein Connexin 26 (Cx26) lead to syndromic deafness, where hearing impairment is associated with skin disease, like in Keratitis Ichthyosis Deafness (KID) syndrome. This condition has been linked to hyperactivity of connexin hemichannels but this has never been demonstrated in cochlear tissue. Moreover, some KID mutants, like Cx26S17F, form hyperactive HCs only when co-expressed with other wild-type connexins. In this work, we evaluated the functional consequences of expressing a KID syndromic mutation, Cx26S17F, in the transgenic mouse cochlea and whether co-expression of Cx26S17F and Cx30 leads to the formation of hyperactive HCs. Indeed, we found that cochlear explants from a constitutive knock-in Cx26S17F mouse or conditional in vitro cochlear expression of Cx26S17F produces hyperactive HCs in supporting cells of the organ of Corti. These conditions also produce loss of hair cells stereocilia. In supporting cells, we found high co-localization between Cx26S17F and Cx30. The functional properties of HCs formed in cells co-expressing Cx26S17F and Cx30 were also studied in oocytes and HeLa cells. Under the recording conditions used in this study Cx26S17F did not form functional HCs and GJCs, but cells co-expressing Cx26S17F and Cx30 present hyperactive HCs insensitive to HCs blockers, Ca2+ and La3+, resulting in more Ca2+ influx and cellular damage. Molecular dynamic analysis of putative heteromeric HC formed by Cx26S17F and Cx30 presents alterations in extracellular Ca2+ binding sites. These results support that in KID syndrome, hyperactive HCs are formed by the interaction between Cx26S17F and Cx30 in supporting cells probably causing damage to hair cells associated to deafness.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Expression of KID syndromic mutation Cx26S17F produces hyperactive hemichannels in supporting cells of the organ of Corti

Abbott

García²,

Villanelo³

et al. 2023

Front. Cell Dev. Biol.

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“…We found that most parasites exhibited only a single homolog, except T. cruzi , T. brucei , and T. rangeli , which possessed five, three, and two, respectively. The expression of two or more homologs could allow the formation of heteromeric channels formed by different subunits as occurs in mammals with Cx26/Cx30 (Naulin et al, 2020), Cx26/Cx46 (Schadzek et al, 2018), Cx43/Cx46 (Hoang et al, 2010) hemichannels among others.…”

Section: Discussionmentioning

confidence: 99%

Unnexins: Homologs of innexin proteins in Trypanosomatidae parasites

Güiza

García

Arriagada

et al. 2021

Journal Cellular Physiology

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Large-pore channels, including those formed by connexin, pannexin, innexin proteins, are part of a broad family of plasma membrane channels found in vertebrates and invertebrates, which share topology features. Despite their relevance in parasitic diseases such as Chagas and malaria, it was unknown whether these large-pore channels are present in unicellular organisms. We identified 14 putative proteins in Trypanosomatidae parasites as presumptive homologs of innexin proteins. All proteins possess the canonical motif of the innexin family, a pentapeptide YYQWV, and 10 of them share a classical membrane topology of large-pore channels. A sequence similarity network analysis confirmed their closeness to innexin proteins. A bioinformatic model showed that a homolog of Trypanosoma cruzi (T. cruzi) could presumptively form a stable octamer channel with a highly positive electrostatic potential in the internal cavities and extracellular entrance due to the notable predominance of residues such as Arg or Lys. In vitro dye uptake assays showed that divalent cations-free solution increases YO-PRO-1 uptake and hyperosmotic stress increases DAPI uptake in epimastigotes of T. cruzi. Those effects were sensitive to probenecid. Furthermore, probenecid reduced the proliferation and transformation of T. cruzi. Moreover, probenecid or carbenoxolone increased the parasite sensitivity to antiparasitic drugs commonly used in therapy against Chagas. Our study suggests the existence of innexin homologs in unicellular organisms, which could be protein subunits of new large-pore channels in unicellular organisms.

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“…It is also possible that other factors recruit connexin proteins, which may lend specific properties (e.g., selectivity or activity) to connexin channels at specific stages. CX26 and CX30 form heteromeric hemichannels, with different selectivities for homomeric versus heteromeric hemichannels [ 19 , 43 , 44 ]. Additionally, the formation of heterotypic channels provides a greater variety in channel properties, including conductance, permeability, and gating, which cannot be obtained with a single connexin [ 45 ].…”

Section: Discussionmentioning

confidence: 99%

Dynamic Spatiotemporal Expression Changes in Connexins of the Developing Primate’s Cochlea

Fujioka

Murayama

Ogawa

et al. 2021

Genes

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Connexins are gap junction components that are essential for acquiring normal hearing ability. Up to 50% of congenital, autosomal-recessive, non-syndromic deafness can be attributed to variants in GJB2, the gene that encodes connexin 26. Gene therapies modifying the expression of connexins are a feasible treatment option for some patients with genetic hearing losses. However, the expression patterns of these proteins in the human fetus are not fully understood due to ethical concerns. Recently, the common marmoset was used as a primate animal model for the human fetus. In this study, we examined the expression patterns of connexin 26 and connexin 30 in the developing cochlea of this primate. Primate-specific spatiotemporal expression changes were revealed, which suggest the existence of primate-specific control of connexin expression patterns and specific functions of these gap junction proteins. Moreover, our results indicate that treatments for connexin-related hearing loss established in rodent models may not be appropriate for human patients, underscoring the importance of testing these treatments in primate models before applying them in human clinical trials.

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Polydisperse molecular architecture of connexin 26/30 heteromeric hemichannels revealed by atomic force microscopy imaging

Cited by 5 publications

References 44 publications

Expression of KID syndromic mutation Cx26S17F produces hyperactive hemichannels in supporting cells of the organ of Corti

Expression of KID syndromic mutation Cx26S17F produces hyperactive hemichannels in supporting cells of the organ of Corti

Unnexins: Homologs of innexin proteins in Trypanosomatidae parasites

Dynamic Spatiotemporal Expression Changes in Connexins of the Developing Primate’s Cochlea

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