Figure 2.
4.1R+17b forms localize and associate with AJ proteins in MDCK cells. A, subcellular localization of 4.1R+17b and E-cadherin in low and normal calcium medium. MDCK cells grown to a confluent state on a coverslip in normal medium (untreated) were switched to low-calcium medium for 20 h (low Ca2+) and returned to normal calcium medium for 8 h (normal Ca2+). Cells were stained with anti-ex17b or anti-E-cadherin Abs and analyzed with Zeiss microscopy. Bar, 10 μm. B, intracellular localizations of 4.1R+17b, β-catenin, E-cadherin, and ZO-1 were examined using its respective antibody in XY and XZ sections and revealed with Zeiss microscopy. XZ sections showed ex17b and E-cadherin or β-catenin, but not ZO-1, co-localized at the AJ. Green, ex17b; red, E-cadherin, β-catenin, or ZO-1. Bar, 10 μm. C, association of 4.1R+17b and AJ proteins in co-immunoprecipitation assays. MDCK cell lysates were precipitated (IP) with an anti-E-cadherin, anti-β-catenin, a control mouse IgG (MIg), anti-HP, anti-ex13, anti-ex17b, or a control rabbit IgG (RIg) Ab. The input extracts and immunoprecipitates were examined by immunoblotting (IB) with its respective antibodies. Molecular mass markers (kDa) are provided at the right margin of each blot.