Cx31 is assembled and trafficked to cell surface by ER-Golgi pathway and degraded by proteasomal or lysosomal pathways.
Gap junctions, consisting of connexins, allow the exchange of small molecules (less than 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 a-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, a-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.
He
Li Qiang
LQ
National Laboratory of Medical Genetics, Central South University, Changsha 410078, China.
Cai
Fang
F
Liu
Yu
Y
Liu
Mu Jun
MJ
Tan
Zhi Ping
ZP
Pan
Qian
Q
Fang
Fai Yan
FY
Liang
De Sheng
DS
Wu
Ling Qian
LQ
Long
Zhi Gao
ZG
Dai
He Ping
HP
Xia
Kun
K
Xia
Jia Hui
JH
Zhang
Zhuo Hua
ZH
D020126:Brefeldin A / Q000494:pharmacology; D017630:Connexins / Q000096:biosynthesis* / Q000235:genetics; D015638:Cytochalasin D / Q000494:pharmacology; D005452:Fluoresceins / Q000378:metabolism; D017629:Gap Junctions / Q000502:physiology*; D006034:Glycyrrhetinic Acid / Q000494:pharmacology; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D015739:Nocodazole / Q000494:pharmacology; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D061988:Proteasome Inhibitors
D017630:Connexins; D005452:Fluoresceins; D061988:Proteasome Inhibitors; C068044:GJB3 protein, human; D020126:Brefeldin A; D015638:Cytochalasin D; D046988:Proteasome Endopeptidase Complex; D006034:Glycyrrhetinic Acid; D015739:Nocodazole; C007740:fluorexon