Figure 6. Yap promotes polyploid cell division via Skp2-mediated FoxO degradation.
(A and B) Immunoblot analysis of FoxO1, FoxO3a and GAPDH (A) and quantitative PCR analysis of FoxO target genes (B) in liver of WT or Mst1/2 DKO mice.
(C) Immunoblot analysis of FoxO1, FoxO3a, Mst1, Mst2, Skp2 and GAPDH in WT, Mst1/2 DKO, Skp2 KO or Mst1f/fMst2f/fSkp2 KO Alb-Cre (TKO) liver lysates.
(D) Immunoblot analysis of p-FoxO1 (S256), p-FoxO1/3 (T24/T32), p-Akt, p-Yap, FoxO1, FoxO3a, Akt, Yap and GAPDH in liver lysates of WT or Mst1/2 DKO mice treated with vehicle control (CTR) or Akt inhibitor MK2206.
(E and F) Immunoblot analysis of the cell fractions (E) or IHC staining (F) of FoxO1 or FoxO3a of liver cells from WT, Mst1/2 DKO, Mst1f/fMst2f/fYAP+/flAlb-Cre (DKO Yap+/−) or Yap Tg mice. Scale bars: 20 µm.
(G) Immunoblot analysis of FoxO1, FoxO3a, α-tubulin or PARP in the cytoplasmic and nuclear fractions of WT, Mst1/2 DKO, Akt1 KO or Mst1/2 Akt1 TKO liver tissues.
(H and I) Immunoassay of the ubiquitination of FoxO1 (detected with anti-Myc) in the lysates of 293T cells expressing various combinations of Myc-tagged ubiquitin, Flag-tagged FoxO1, HA-tagged Yap and HA-tagged Skp2 (H) with or without Akt inhibitor MK2206 treatment
(J) Quantification of BrdU or TUNEL-positive cells in the liver sections from Mst1/2 DKO mice infected with Ad-GFP or Ad-Foxo1(S256A) (n = 3).
(K and L) The representative liver sizes and liver-to-body weight ratios of Mst1/2 DKO infected with adenovirus Ad-GFP or Ad-FoxO1(S256A) (n = 5) (K) or Skp2 KO mice infected with adenovirus Ad-GFP, Ad-FoxO1(S256A), Ad-Skp2(KLKL) or Ad-FoxO1(S256A) plus Ad-Skp2(KLKL) (n = 5) (L).
(M) A proposed working model for Yap promoting polyploid cell division via Skp2-mediated FoxO degradation.
Data were assessed by Student’s t-test and represented as mean ± SD *p < 0.05, **p < 0.01, ***p < 0.001 compared between the indicated groups.
See also Figure S6.