Figure 7.

The interaction of Xenopus occludin with chicken occludin in vivo assayed by immunoprecipitation. The embryos were microinjected with the most COOH-terminally truncated occludin mRNA (266; a and b, left lanes) or water (C; a and b, right lanes) and incubated at room temperature for 10 h. After homogenization, the samples were centrifuged at 100,000 g for 30 min at 4°C. The membrane pellet was solubilized in modified RIPA buffer (see Materials and Methods) and centrifuged at 100,000 g for 1 h at 4°C. The resulting supernatant was immunoprecipitated with anti-occludin 11350 (a) or anti-FLAG M2 (b) overnight at 4°C. Immunoprecipitates were immunoblotted with anti-FLAG M2 (a) or anti-occludin 11350 (b). A specific band corresponding to the chicken truncated occludin (compare with Fig. 2 b, lane 266) was present in a (left lane) after immunoprecipitated with anti-occludin 11350. The Xenopus full-length occludin was observed at b (left lane) after immunoprecipitated with anti–FLAG M2. The two thick bands in b were IgG heavy chain.