PKD1 is required for curcumin induced enrichment of β-catenin on the membrane.

<p>A). Silencing of PKD1 by PKD1 specific siRNA. C4-2 cells were transfected for 48 h with 25 nM control siRNA or PKD1 siRNA, lysed and immunoblotted for PKD1 and β-actin using specific antibodies. Quantitation of protein band intensities was performed by densitometric analysis. The PKD1 levels was normalized to β-actin levels and graphed. AU- arbitrary units. Immunoblotting shows over 95% suppression of PKD1 expression on transfection with PKD1 specific siRNA (lane 2) compared to control siRNA-transfected cells (lane 1). B). Suppression of PKD1 inhibits enrichment of membrane β-catenin levels. C4-2 cells were cultured on coverslips overnight. The cells were first transfected with either control siRNA (A1–D1; A2–D2) or PKD1 silencing siRNA (A3–D3; A4–D4) for 24 h, followed by treatment with vehicle control (DMSO) (A1–D1; A3–D3) or curcumin (20 µM) (A2–D2; A4–D4) for 1 h. The cells were immunostained for β-catenin (red) or PKD1 (green) and the nucleus was counter stained with DAPI (blue). Higher β–catenin staining was observed on the cell surface of control siRNA cells at 1 h of curcumin treatment (A2) compared to vehicle treatment (A1). However, siRNA mediated silencing of PKD1 (B3, B4) inhibited curcumin mediated enrichment of membrane β–catenin staining on the cell surface (A4 vs A3 and A2). Original Magnifications 600× with 2× zoom.