Figure 8.

Knockdown of Rap1 inhibits junctional localization of cadherin during junctional reassembly after calcium switch. ARPE-19 cells were plated onto coverslips at confluent density, and then treated with negative control or Rap1 RNAi for 48 hours. Cells were then treated with EGTA for 30 minutes to disrupt cell-cell junctions, followed by washout and readdition of calcium-containing media for indicated times. Coverslips were fixed and processed for immunofluorescent detection with a pan-cadherin antibody to visualize cell-cell junctions. EGTA treatment completely disrupts the cell monolayer of both control and Rap1 knockdown. On washout of EGTA, control cell monolayers regain junctional cadherin localization and intact monolayer appearance faster than Rap1 knockdown cells.