Figure 3.

YAP and TAZ activity is decreased upon NUP205 downregulation. (A) Co-immunoprecipitation of endogenous YAP shows a decrease in interaction with TEAD1 in HEK293T cells transfected with siRNA NUP205 compared with control. IgG is used as negative control for the IP. Quantification of relative TEAD1 pulldown normalized to TEAD1 lysates strength and further normalization to strength of YAP-immunoprecipitation (n = 4). (B) qPCR shows decrease of relative mRNA expression of YAP/TAZ target genes in HEK293T cells transfected with siRNA NUP205 compared with control (n = 5). ^*Significant decrease by siRNA NUP205 (unpaired t-test; P < 0.05). (C) Proliferation rate of cultured podocytes by live-cell imaging over the period of 24 h. Knockdown of Nup205 or Yap inhibited the proliferation activity as compared with control cells. Representative images for cell confluency after 16 h. (D) Cell death assessed by live-cell imaging over the period of 24 h. Dead cells marked with DiYO-1. Basal cell death (untreated cells; dashed lines depicted in both left and right panels) and after treatment with either 100 μM H₂O₂ (lines, left panel) or 1 μg/ml ADR (lines, right panel). Lack of Nup205 or Yap resulted in decreased resistance to oxidative stress, when compared with scrambled controls. Live-cell images were captured every 2 h (n = 3–4 plates). Data are reported as mean ± SEM.