A pore locus in the E1 domain differentially regulates Cx26 and Cx30 hemichannel function

Area of Science:

• Cellular biology
• Biophysics
• Otolaryngology

Background:

• Connexins (Cxs) form gap junction (GJ) and hemichannels crucial for cellular communication.
• Cx26 and Cx30 are prevalent in the cochlea, playing roles in hearing.
• The E1 loop's parahelix is implicated in channel gating.

Purpose of the Study:

• Investigate the role of the Cx26/Cx30 E1 parahelix in hemichannel function.
• Determine the impact of the Ala/Glu difference at position 49.
• Analyze interactions at positions 49 and 50 in hemichannel and GJ channel activity.

Main Methods:

• Cysteine-scanning and mutational analyses of Cx26 and Cx30.
• Co-expression studies involving wild-type and mutant connexins.
• Electrophysiological recordings of hemichannel and GJ channel currents.
• Analysis of cochlear explants for calcium wave propagation.

Main Results:

• Position 49 is pore-lining and interacts with Asp50, modulating hemichannel function.
• Charged residues at positions 49/50 (as in Cx30) dampen hemichannel activity.
• The Cx26 D50N mutation, linked to deafness, rescues hemichannel function in Cx30 co-expression.
• Cysteine substitutions at position 49 inhibit hemichannels via disulfide or metal coordination.
• GJ channels are unaffected by these mutations, unlike hemichannels.
• Cx30 expression correlates with reduced calcium wave propagation in cochlear explants.

Conclusions:

• A specific pore locus in the E1 loop of Cx26 and Cx30 significantly impacts hemichannel function.
• Interactions between positions 49 and 50 are critical for hemichannel gating and activity.
• These findings offer new insights into connexin roles in cochlear physiology and disease.