Heteromeric, but Not Homomeric, Connexin Channels Are Selectively Permeable to Inositol Phosphates

Abstract

Previous work has shown that channels formed by both connexin (Cx)26 and Cx32 (heteromeric Cx26/Cx32 hemichannels) are selectively permeable to cAMP and cGMP. To further investigate differential connexin channel permeability among second messengers, and the influence of connexin channel composition on the selectivity, the permeability of inositol phosphates with one to four phosphate groups through homomeric Cx26, homomeric Cx32, and heteromeric Cx26/Cx32 channels was examined. Connexin channels were purified from transfected HeLa cells and from rat, mouse, and guinea pig livers, resulting in channels with a broad range of Cx26/ Cx32 aggregate ratios. Permeability to inositol phosphates was assessed by flux through reconstituted channels. Surprisingly, myoinositol and all inositol phosphates tested were permeable through homomeric Cx32 and homomeric Cx26 channels. Even more surprising, heteromeric Cx26/Cx32 channels showed striking differences in permeability among inositol phosphates with three or four phosphate groups and among isomers of inositol triphosphate. Thus, heteromeric channels are selectively permeable among inositol phosphates, whereas the corresponding homomeric channels are not. There was no discernible difference in the permeability of channels with similar Cx26/Cx32 ratios purified from native and heterologous sources. The molecular selectivity of heteromeric channels among three inositol triphosphates could not be accounted for by simple connexin isoform stoichiometry distributions and therefore may depend on specific isoform radial arrangements within the hexameric channels. Dynamic regulation of channel composition in vivo may effectively and efficiently modulate intercellular signaling by inositol phosphates.Connexin channels, which compose most vertebrate gap junctions, mediate direct intercellular movement of ions and molecules. There are ϳ20 isoforms of connexin protein, each forming channels with distinct functional properties. Every known functional deletion of a connexin isoform produces a distinct pathology, and genetic replacement of one connexin (Cx) 2 by another ("knock in") fails to fully compensate (1-3). Pathologies that arise from altered connexin channel function must arise from abnormal molecular movement through connexin channels, whether in magnitude, regulation, or molecular specificity (reviewed in Ref. 4).Gap junction channels form by end-to-end interaction of hemichannels, each consisting of six connexin monomers. Hemichannels are either homomeric (composed of a single connexin isoform) or heteromeric (more than one isoform). Dramatic and surprising degrees of ionic and molecular permselectivity have been observed for homomeric channels (5-13). However, most cells express more than one connexin, and heteromeric connexin channels are common in vivo (14 -18). Heteromeric mixing of different connexin isoforms, producing variation in channel stoichiometry and/or arrangements of isoforms within the hemichannels, may allow cells to dynamically regulate their intercellular co...

Connexin channel permeability to cytoplasmic molecules

“…The work with reconstituted heteromeric Cx26/Cx32 hemichannels shows that some of these channels were permeable to IP3 and some were not, whereas all the inositol triphosphates tested permeated the corresponding homomeric hemichannels (Ayad et al, 2006). This suggests that heteromeric channels can have higher degrees of selectivity than homomeric channels.…”

“…In addition, the fact that some of the Cx26/Cx32 channels are impermeable to some inositol triphosphates and permeable to others (Ayad et al, 2006) makes clear that pore diameter and permeant charge are not the key determinants of the selective permeability.…”

“…aspartate (Ye et al, 2003) 3a 3c 3e 5 glutamate (Ye et al, 2003) 3a 3c 3e 5 26/32h cAMP (Bevans et al, 1998;Ayad et al, 2006) 3f (not permeable through all the heteromeric channels)…”

“…IP3 (Ayad et al, 2006) 3f (not permeable through all the heteromeric channels; differential permeability among inositol triphosphate isoforms) 26j cAMP (Bedner et al, 2003;Bedner et al, 2006) 1a IP3 1b 2…”

ATP release through connexin hemichannels and gap junction transfer of second messengers propagate Ca 2+ signals across the inner ear

“…Interestingly, ICS propagation across heteromeric GJ channels consisting of Cx26 and Cx30 is reported to be faster than across their homomerically assembled counterparts (30). Furthermore, heteromeric mixing of different connexin isoforms, producing variation in channel stoichiometry and/or arrangements of isoforms within the hemichannels, may allow cells to regulate their intercellular communication properties, including molecular selectivity to inositol phosphates (31). Permeability to IP 3 is impaired in a deafness mutant of Cx26 (9).…”