Supramolecular dynamics of gap junctions.
During the life cycle of a membrane protein its molecular structure may change and for aggregated proteins this process may be observed on the supramolecular level. Here we demonstrate that this is the case for gap junction channels which maintain cell-cell communication. Freshly synthesized connexins are integrated as hexamers (connexons) into the plasma membrane where they form plaques after pairing with connexons of an attached cell. We inhibited protein trafficking by applying the fungal metabolite brefeldin A (BFA), quantified cell-cell coupling by calcein transfer and fluorescence-activated flow cytometry, and examined the degradation and formation of gap junction plaques by indirect immunofluorescence and immunogold labeling. Under control conditions 50% of the detected plaques were ubiquitylated and less than 10% showed a two-dimensional crystalline packing. One hour after BFA reversal about 60% of the plaques were crystalline and ubiquitylation dropped to 14%. Label for ubiquitin was predominantly found on non-crystalline plaques. We, therefore, conclude that newly formed gap junction plaques are of crystalline morphology which changes to a pleomorphic structure when individual channels are modified during their aging process. This dynamic in plaque morphology correlates with channel inactivation and plaque ubiquitylation.
Rütz
M L
ML
Biologisches Institut, Abeteilung Biophysik, Universität Stuttgart, Germany.
Hülser
D F
DF
D000818:Animals; D018031:Connexin 43 / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D017629:Gap Junctions / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D014407:Tumor Cells, Cultured
D018031:Connexin 43