A, Tight monolayers of MDCK cells were grown in the absence (lanes 1–4) or presence of swainsonine (lanes 5–8) as described in the legend to Fig. 5. Cell monolayers were biotinylated from the basolateral side as described in Experimental procedures. As indicated, right before lysis, cells were incubated either with PBS (lanes 1 and 5), or with 0.25% (lanes 2 and 6), or with 0.5% (lanes 3 and 7) or with 0.75% Triton X-100 (lanes 4 and 8) in PBS for 15 min and rinsed with PBS. The gel mobility and detergent resistance of both E-cadherin and the Na,K-ATPase β1 subunit increases in the presence of swainsonine as detected by a Western blot analysis.
B–C, Quantification of the results showing that swainsonine significantly increases the resistance of the Na,K-ATPase β1 subunit (B) and E-cadherin (C) to Triton X-100 extraction.
Error bars, ± s.d. (n=3). Significant difference with untreated cells: * P<0.05, ** - P<0.001, Student’s t-test.