Small synthetic peptides homologous to segments of the first external loop of occludin impair tight junction resealing.
This study shows that resealing of opened tight junctions (TJs) is impaired by interaction with oligopeptides homologous to the external domain of chick occludin. The experiments were carried out with confluent A6 cell monolayers grown on collagen supports under stable transepithelial electrical resistance (TER). The monolayers were bathed on the apical side with a 75 mm KCl solution and on the basolateral side by NaCl-Ringer's solution. TJ opening was induced by basolateral Ca2+ removal and was characterized by a marked drop of TER. The reintroduction of Ca2+ triggered junction resealing as indicated by an elevation of TER to control values. Custom-made peptides SNYYGSGLSY (corresponding to the residues 100 to 109) and SNYYGSGLS (residues 100 to 108), homologous to segments of the first external loop of chick occludin molecule, impaired junction resealing when the peptides were included in the apical bathing fluid (concentrations in the range of 0.5 to 1.5 mg/ml). Peptide removal from the apical solution usually triggered a slow recovery of TER, indicating a slow recovery of the TJ seal. Changes in localization of ZO-1, a cytoplasmic protein that underlies the membrane at the TJs, were evaluated immunocytochemically following Ca2+ removal and reintroduction. The presence or absence of the oligopeptides showed no influence on the pattern of change of ZO-1 localization. These observations support the hypothesis that the TJ seal results from the interaction of specific homologous segments of occludin on the surface of adjacent cells. Additionally, our results show that small peptides homologous to segments of the occludin first external loop can be used as specific reagents to manipulate the permeability of tight junctions.
Lacaz-Vieira
F
F
Departamento de Fisiologia e Biofísica, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP, Brazil.
Jaeger
M M
MM
Farshori
P
P
Kachar
B
B
D000818:Animals; D002021:Buffers; D002118:Calcium / Q000494:pharmacology; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000187:drug effects*; D016764:Cell Polarity; D002645:Chickens; D007365:Intercellular Junctions / Q000187:drug effects*; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D062793:Occludin; D010446:Peptide Fragments / Q000494:pharmacology*; D010750:Phosphoproteins / Q000378:metabolism; D062826:Zonula Occludens-1 Protein
D002021:Buffers; D008565:Membrane Proteins; D062793:Occludin; D010446:Peptide Fragments; D010750:Phosphoproteins; D062826:Zonula Occludens-1 Protein; D002118:Calcium