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. 2008 Dec 29;106(1):61–66. doi: 10.1073/pnas.0802741106

Fig. 5.

Fig. 5.

PKCη phosphorylates occludin on specific Thr residues. (A) Alignment of amino acid sequence of the C-terminal region of occludin from different species. Conserved Thr residues are highlighted. The Thr residues in human occludin selected for mutation are identified by arrows. (B) GST-Occludin-C and its Thr-mutants (p44) were prepared and incubated with PKCη in the presence or absence of ATP. Reaction mixtures were immunoblotted to p-Thr and GST. (C) Densitometric analysis of p-Thr bands in experiments described in B. (D and E) GFP-Occludin (p90) and its Thr-mutants were expressed in MDCK (D) and Caco-2 (E) cells. Anti-GFP immunocomplexes were immunoblotted for p-Thr and GFP. Densitometric analysis was performed to evaluate the p-Thr densities. T3A refers to occludin in which T403, T404, and T438 were mutated and T5A to occludin with all 5 threonines mutated. In all experiments, values are mean ± SEM (n = 3). Asterisks indicate the values that are significantly different (P < 0.05) from corresponding value for vector-transfected cells.