Figure 5.

Alteration of apical protein expression and sorting accompanies deciliation. (A) Polarized cultures of MDCK II cells were treated as described in Figure 3, and secretion of gp80/clusterin was measured as described in Materials and Methods. Note that apical secretion of this protein (control) becomes randomized after deciliation (+ CH), but then returns to normal after regrowth of cilia (washout). n = 4 filters per condition; error bars, SEM. (B) Left, polarized MDCK II cells were fixed and labeled with antibodies to gp114/CEACAM. En face (top) and orthogonal (bottom) images are shown. Bar, 5 μm. Right, triplicate cultures of polarized MDCK II cells were biotinylated apically or basolaterally. Biotinylated proteins were precipitated on streptavidin agarose, resolved by SDS-PAGE, and immunoblotted for gp114/CEACAM. Levels of protein recovered were quantified by phosphorimaging, and results are presented as % of total surface-labeled gp114/CEACAM. Note that apical polarity of gp114/CEACAM is lost in deciliated cultures. n = 3; error bars, SEM. (C) Left, polarized MDCK II cells were fixed and labeled with antibodies to gp135/podocalyxin. En face (top) and orthogonal (bottom) images are shown. Bar, 10 μm. Right, surface biotinylation assay was performed as in B. Note that expression level of gp135/podocalyxin is greatly decreased in deciliated cultures.