Fig 2: Effects of the estrogen receptor antagonist (ICI 182, 780, ICI) on the levels of YAP and phospho-YAP (YAP S127) in the uterus of OVX mice. (A) Cross-sections of uterus from sesame oil (oil; control), E2 only, and/or ICI+E2-treated OVX mice were stained with rabbit polyclonal antibodies against YAP or phospho-YAP (YAP S127). DAPI is shown in blue. LE, luminal epithelium; GE, glandular epithelium. Scale bar: 25 µm. (B,C) Relative quantitation of YAP and phospho-YAP (YAP S127) of immunofluorescence staining was analyzed using LAS AF Life software.

Fig 3: STK25 regulates Hippo activation in response to loss of cytoskeletal tension. a Immunoblot and quantitation of phosphorylated YAP levels following treatment with 10 µM DCB in HEK293A cells transfected with the indicated siRNA (n = 6; ***p < 0.001, unpaired t-test). b Global phosphorylation status of YAP was assessed using Phos-tag gel electrophoresis following treatment with 10 µM DCB in HEK293A cells transfected with either control siRNA or STK25 siRNA. Shifted bands indicate degrees of YAP phosphorylation. c TAZ phosphorylation status was assessed using Phos-tag gel electrophoresis following treatment with 10 µM DCB in HEK293A cells transfected with either control siRNA or STK25 siRNA. d Immunoblot and quantitation of phosphorylated YAP levels following treatment with 10 µM DCB in either control HEK293A stably expressing Cas9 and a non-targeting sgRNA or STK25 KO 293A stably expressing Cas9 together with either sgRNA 1 (Clone 1) or sgRNA 2 (Clone 2) targeting STK25 (n = 4; *p < 0.05, one-way ANOVA with Dunnett’s post-hoc analysis). e Immunoblot and quantitation of phosphorylated YAP levels in control 293A cells and STK25 KO 293A cells transfected with either Vector, Cas9-resistant FLAG-STK25-WT, or Cas9-resistant FLAG-STK25-KD. Global levels of YAP phosphorylation in these samples were also assessed using Phos-tag gel electrophoresis. Quantitation corresponds to levels of phosphorylated YAP as measured via phos-tag electrophoresis (n = 4; ***p < 0.001, ****p < 0.0001; One-way ANOVA with Dunnett’s post-hoc analysis). All data are presented as mean ± SEM

Fig 4: Empagliflozin decreased Yes-associated protein (YAP) expression in LX-2 cells. (a) The representative protein expression of YAP and phospho-YAP was determined using Western blot. (b) Quantitation and normalisation of phospho-YAP to total YAP or total YAP to β-actin from a. (c) The relative expression of cyr61, cTGF, and AREG was determined via RT-PCR. Data are expressed as means ± standard deviation of three independent experiments. * p < 0.05, ** p < 0.01, and *** p < 0.001 compared to the control.

Fig 5: Tubular epithelial YAP activation associated with macrophage infiltration and renal MCP-1 upregulation.A Representative image of double immunofluorescent staining with YAP and F4/80 in Sham and IRI-28d kidneys. B YAP-activated tubules and F4/80-positive infiltrates were calculated from 55 randomly captured images at IRI-28d kidneys. Correlation between YAP-activated tubules and F4/80-positive infiltrates was analyzed (n = 55). Pearson correlation R value and P value are indicated. C Gene expression analysis for chemokine members including CCL and CXCL family in Mst1/2 knockout mice. Gene Expression Omnibus (GEO) datasets supply the RNA-sequencing data of GES95463. D Regulatory elements of the genes encoding MCP-1 from various mammals were reported by Kim et al.[20]. The TEAD-binding element (red bar) is located 100–400 bp upstream of the transcription start site of MCP-1. TSS transcription start site. E Representative western blot analysis of MCP-1 protein expression in kidney lysates from mice 14 and 28 days after IRI (n = 6 mice per group). Sham-operated mice was used as a control (n = 3 mice). F ELISA analysis of MCP-1 levels in serum. G, H Quantitative polymerase chain reaction analysis of MCP-1 (Ccl2) and CCR2 (Ccr2) mRNA expression in kidneys. AU arbitrary units. Scale bar = 50 µm in all images of A. All values are means ± SEM. *p < 0.05, **p < 0.01, and #p < 0.001 defined as significant. ns not significant.