Cx31 is assembled and trafficked to cell surface by ER-Golgi pathway and degraded by proteasomal or lysosomal pathways

Abstract

Gap junctions, consisting of connexins, allow the exchange of small molecules (<1 kD) between adjacent cells, thus providing a mechanism for synchronizing the responses of groups of cells to environmental stimuli. Connexin 31 is a member of the connexin family. Mutations on connexin 31 are associated with erythrokeratodermia variabilis, hearing impairment and peripheral neuropathy. However, the pathological mechanism for connexin 31 mutants in these diseases are still unknown. In this study, we analyzed the assembly, trafficking and metabolism of connexin 31 in HeLa cells stably expressing connexin 31. Calcein transfer assay showed that calcein transfer was inhibited when cells were treated with Brefeldin A or cytochalasin D, but not when treated with nocodazole or α-glycyrrhetinic acid, suggesting that Golgi apparatus and actin filaments, but not microtubules, are crucial to the trafficking and assembly of connexin 31, as well as the formation of gap junction intercellular communication by connexin 31. Additionally, α-glycyrrhetinic acid did not effectively inhibit gap junctional intercellular communication formed by connexin 31. Pulse-chase assay revealed that connexin 31 had a half-life of about 6 h. Moreover, Western blotting and fluorescent staining demonstrated that in HeLa cells stably expressing connexin 31, the amount of connexin 31 was significantly increased after these cells were treated with proteasomal or lysosomal inhibitors. These findings indicate that connexin 31 was rapidly renewed, and possibly degraded by both proteasomal and lysosomal pathways.

Connexin Expression and Functional Analysis of Gap Junctional Communication in Mouse Embryonic Stem Cells

“…Therefore, we conclude that the gap junction inhibitor acts on Cx43 but not on Cx45 or Cx31. Similar findings were published by He et al [29] with regard to Cx31.…”

Pannexins and gap junction protein diversity

“…In particular, experiments using transient expression of this protein in cultured cells [27,62,64,93,103] showed abundant accumulation in the Golgi apparatus and the ER. One feasible explanation for the ER and Golgi labeling implies that posttranslational modification and assembly of pannexons share the same route demonstrated for connexins [143,144]. The ERbound Panx1 was, therefore, suggested to be either a pool of unprocessed precursor proteins [64,103] or assembled functional pannexons that serve as ER calcium release channels, thus providing a conduit for the enigmatic Ca 2+ leak across the ER membrane [62].…”

Pannexins, distant relatives of the connexin family with specific cellular functions?

“…(140,141) The ER-bound Panx1 could be either a pool of unprocessed precursor proteins (121,138) or assembled functional pannexons that serve as ER-Ca 2þ -release channels, thereby facilitating Ca 2þ leakage from the ER. (101) Panx trafficking has been investigated by differences between tagged and untagged Panxs and by treating the cells with brefeldin A (BFA), (32,33) which promotes Golgi breakdown.…”