Fig 2: Strictinin modulates ROR1-mediated AKT-GSK3β signaling in DU145 cells. (A) Immunoblot on DU145 cells treated with control, 250 µmol/L strictinin for 24 h, or 250 µmol/L strictinin for 48 h. ROR1 is visualized; (B) quantification of ROR1 protein band normalized to GAPDH. Data are represented as the mean ± SEM from three independent western blot experiments; (C) immunoblot on DU145 cells treated with control, 250 µmol/L strictinin for 24 h, or 250 µmol/L strictinin for 48 h. AKT, p-AKT-ser473, GSK3β, p-GSK3β-ser9 visualized; (D) quantification of p-AKT/AKT and p-GSK3β/GSK3β protein bands normalized to GAPDH. Data are represented as the mean ± SEM from three independent western blot experiments. N = 3. ** P value < 0.01; * P value < 0.05

Fig 3: Higher level of Rif expression is associated with poorer prognosis in patients with LUAD and its aggressiveness.A, Kaplan–Meier survival analysis of Rif high and low expression groups in patients with LUAD. p values are obtained from the log-rank test. B, expression levels of Rif in LUAD were analyzed in correlation with the indicated classified tumor grades. Statistical significance was analyzed using t test. C, immunohistochemical analysis of Ror1 and Rif on tissue microarray of LUAD. Representative images of double-positive (Rif+/Ror1+) and single-positive (Rif+/Ror1-or Rif-/Ror1+) specimens and a summary of the analyses (n = 42) are shown. The scale bars represent 200 μm. LUAD, lung adenocarcinoma; Rif, Rho in filopodia.

Fig 4: Ror and Rif induce formation of dorsal filopodia and interact with each other.A and B, representative images of HeLaS3 cells expressing the indicated fluorescence proteins on a 2D surface. Phalloidin staining was performed to visualize F-actin (A). Serial optical confocal z sections are stacked (z-stack), and the xz images sectioned along the white dotted line in the z-stack images are shown. Insets show magnified images of boxed regions. Images are representative of at least three independent experiments. The scale bars represent 10 μm (main images) and 2 μm (magnified images). C, coimmunoprecipitation assay showing the association of Flag-Rif with Ror1-mCherry (mCh) and Ror2-mCh in 293T cells. Whole-cell lysates (WCL) from 293T cells expressing the indicated proteins were subjected to immunoprecipitation (IP) with anti-RFP antibody (that recognizes mCherry), followed by Western blotting. Blots are representative of three independent experiments. Rif, Rho in filopodia

Fig 5: Survival Analysis. (A) Survival analysis of all PDAC patients with and without peritoneal carcinomatosis (PDAC (PER+) + PDAC (PER-)). p=0.0482, n=27. (B) Survival analysis of only PDAC (PER-) patients without peritoneal carcinomatosis. p=0.1699, n=18. Blue: High expression of exo-ROR1, Black: Low expression of exo-ROR1. (C) Survival analysis according to ROR1 mRNA in tumor tissue. p=0.8414, n=25. Blue: High expression of ROR1 mRNA, Black: Low expression of ROR1 mRNA.