Multiple Protein Interactions Involving Proposed Extracellular Loop Domains of the Tight Junction Protein Occludin

Abstract

Occludin is a tetraspan integral membrane protein in epithelial and endothelial tight junction (TJ) structures that is projected to have two extracellular loops. We have used peptides emulating central regions of human occludin's first and second loops, termed O-A:101-121 and O-B:210-228, respectively, to examine potential molecular interactions between these two regions of occludin and other TJ proteins. A superficial biophysical assessment of A:101-121 and O-B:210-228 showed them to have dissimilar solution conformation characteristics. Although O-A:101-121 failed to strongly interact with protein components of the human epithelial intestinal cell line T84, O-B:210-228 selectively associated with occludin, claudin-one and the junctional adhesion molecule (JAM)-A. Further, the presence of O-B:210-228, but not O-A:101-121, impeded the recovery of functional TJ structures. A scrambled peptide sequences of O-B:210-228 failed to influence TJ assembly. These studies demonstrate distinct properties for these two extracellular segments of the occludin protein and provide an improved understanding of how specific domains of occludin may interact with proteins present at TJ structures.

Regulation of Heterotypic Claudin Compatibility

“…The compatibility profile for claudin-1, claudin-3, and claudin-5 has also been observed for airway epithelia (14). Although occludin-null mice form functional tight junctions (17,26), the ability of occludin EL peptides to disrupt epithelial barrier function suggests a role for occludin in regulating tight junctions (18,19,27). At present it is unclear whether heterotypic occludin-claudin interactions contribute to the ability of occludin to modulate barrier function.…”

Junctional proteins of the blood-brain barrier: New insights into function and dysfunction

“…1). 2 The TJ transmembrane proteins include the integral membrane proteins, occludin and claudins (for example, claudin-5, ¡3, ¡12, ¡1) and an IgG type of protein, junctional adhesion molecule (JAM) -A, -B and -C. 3 Claudins, the primary sealing components of TJs, are a family of transmembrane proteins (20)(21)(22)(23)(24), with more than 24 isoforms identified. 4 Most claudins show tissue specific expression and their presence in cells is characterized as constitutive and/or inducible.…”

Wound healing revised: A novel reepithelialization mechanism revealed by in vitro and in silico models

“…For 2D experiments, HaCaTs were cultured using culture slides until they reached a confluency of 70-80%. Cells were washed with HBSS / , and tight junctions were disrupted by using a Ca 2+ switch protocol according to Nusrat et al (2005). Peptides having a concentration of 200 µM were applied, and cells were incubated at 37°C for 5 and 24 h. Organotypic cultures were wounded, and 4 µl peptide solution having a concentration of 400 µM was applied every hour directly into the wound for a period of 6 h. Fig.…”