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. 2013 Dec;183(6):1945–1959. doi: 10.1016/j.ajpath.2013.08.026

Figure 6.

Figure 6

Overexpression of PKCλ/ι rescues the phenotype of PKCλ/ι−/− podocytes. A: Real-time PCR analysis of PKCλ/ι+/+ and PKCλ/ι−/− podocytes infected with adenoviral PKCι-GFP or GFP as control. Transduction with PKCι-GFP rescued the mRNA expression of PKCι and reduced the level of Def-6 mRNA in PKCλ/ι−/− podocytes. B: Western blot analysis demonstrated overexpression of PKCι-GFP or GFP control virus. C: Phalloidin stainings of PKCλ/ι+/+ and PKCλ/ι−/− podocytes transduced with adenoviral PKCι-GFP or GFP as control. Transduction with PKCι-GFP rescued the actin cytoskeletal phenotype of the PKCλ/ι−/− podocytes. Images were obtained using the Leica DMBL microscope (×20 magnification). Scale bar = 50 μm (applies to all panels). D: Measurement of cell area and number of focal adhesions of vinculin-stained transduced podocytes. ∗∗P < 0.01, ∗∗∗P < 0.001. E: Representative vinculin staining after virus transduction. Scale bar = 50 µm (applies to all panels). F: Overexpression of Def-6 in podocytes. In transfected podocytes, Def-6 FL GFP localizes to the cellular edges. Overexpression of constitutively active Def-6 DHL leads to enhanced membranous localization (boxed areas in GFP views and arrows in Detail views). As control, podocytes were transfected with pEGFP-N1. G: Transduced PKCλ/ι+/+ and PKCλ/ι−/− podocytes stained with an antibody against Def-6. Overexpression of PKCι reversed the enhanced membrane localization of Def-6 in PKCλ/ι−/− podocytes (boxed areas in GFP views and arrows in middle Detail view).