The SARS Coronavirus E Protein Interacts with PALS1 and Alters Tight Junction Formation and Epithelial Morphogenesis

Teoh, Kim-Tat; Siu, Yu-Lam; Chan, Wing-Lim; Schlüter, Marc A.; Liu, Chia-Jen; Peiris, J. S. Malik; Bruzzone, Roberto; Margolis, Benjamin; Nal, Béatrice

Our data also showed that expression of E (wt), but not E (ΔPBM), delayed TJ formation in MDCKII cells in calcium switch assays (Figure 6). This is well illustrated by immunofluorescence and confocal microscopy analysis of these cells (Figures 7 and 8). Indeed, two hours post-calcium switch, E (wt) expressing cells present a strong defect of polarity with a mis-location of polarity markers, whereas control cells and E (ΔPBM) expressing cells are polarized, with TJ (ZO-1, PALS1), AJ (E-cadherin) and apical (GP135) markers correctly localized. These data indicate that E expression alters TJ formation in a PBM-dependent manner, and affects establishment of polarity. A significant number of E (wt) expressing cells were round with presence of PALS1 in the cytoplasm, occasionally found associated with E in the Golgi region. This was generally observed for cells with higher expression levels of E. However, TJ could form, although with a delay of several hours, suggesting that the kinetics of trafficking of PALS1 was affected but not the process of TJ formation. At 120 h post-calcium switch, PALS1 was present at cell–cell contacts for both E (wt) and E (ΔPBM) expressing cells, witch are most likely back to a steady state with low TER values, common for this MDCKII cell line (Figure 8, panels g and h). However, round cells were still present in E (wt) expressing cells. Conversely, E (ΔPBM) expressing cells showed enhanced TER values, which indicated a rapid formation of junctions. E (ΔPBM) was not present in the Golgi region but was rather diffuse in the apical region of the cytosol. PALS1 was not mislocalized and did not colocalize with this truncated form of E. Therefore, we conclude that SARS-CoV E alters PALS1 distribution in monolayers of MDCKII cells and, as a consequence, disturbs TJ and polarity in a PBM-dependent manner.