Redox-sensitivity of the dimerization of occludin.
Occludin is a self-associating transmembrane tight junction protein affected in oxidative stress. However, its function is unknown. The cytosolic C-terminal tail contains a coiled coil-domain forming dimers contributing to the self-association. Studying the hypothesis that the self-association is redox-sensitive, we found that the dimerization of the domain depended on the sulfhydryl concentration of the environment in low-millimolar range. Under physiological conditions, monomers and dimers were detected. Masking the sulfhydryl residues in the domain prevented the dimerization but affected neither its helical structure nor cylindric shape. Incubation of cell extracts containing full-length occludin with sulfhydryl reagents prevented the dimerization; a cysteine/alanine exchange mutant also did not show dimer formation. This demonstrates, for the first time, that disulfide bridge formation of the domain is involved in the occludin dimerization. It is concluded that the redox-dependent dimerization of occludin may play a regulatory role in the tight junction assembly under physiological and pathological conditions.
Walter
J K
JK
Leibniz-Institut für Molekulare Pharmakologie Berlin, FMP, Robert-Rössle-Str. 10, 13125, Berlin-Buch, Germany.
Castro
V
V
Voss
M
M
Gast
K
K
Rueckert
C
C
Piontek
J
J
Blasig
Ingolf E
IE
D000818:Animals; D002478:Cells, Cultured; D004220:Disulfides / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D062793:Occludin; D010084:Oxidation-Reduction / Q000187:drug effects; D054730:Protein Interaction Domains and Motifs / Q000187:drug effects; D055503:Protein Multimerization / Q000187:drug effects / Q000502:physiology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013329:Structure-Activity Relationship; D019108:Tight Junctions / Q000187:drug effects / Q000378:metabolism
D004220:Disulfides; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; C568986:Ocln protein, mouse; D012967:Sodium Dodecyl Sulfate; D004229:Dithiothreitol