Fig. 4

COMP activates Notch3 receptor and interacts with Jagged1 ligand. Analyses of the mRNA expression levels of NOTCH1, NOTCH2, NOTCH3, and JAG1 in A SKOV3 and B OAW42 cells treated with different concentrations of COMP (20 and 50 μg/ml) or BSA (50 μg/ml) as a control by RT-qPCR. The p-value was calculated by two-way ANOVA followed by Tukey’s post-test. C The activation of Notch receptors in response to COMP (20 and 50 μg/ml) was assessed by western blot analyses in SKOV3 and OAW42 cells. PBS-treated and BSA (50 μg/ml)-treated cells were used as negative controls. D, E The protein expression levels of NICD3 were quantified by densitometry and normalized to GAPDH. One-way ANOVA followed by Dunnett’s post-test was used for statistical analyses. F The colocalization of Notch3 receptor and its ligand Jagged1 was evaluated in response to COMP (0.25 mg/ml) by PLA assay in G SKOV3 and H OAW42 cells (Blue areas: DAPI-stained nuclei, white dots: Notch3-Jagged1 co-expression). The p-value was calculated by unpaired t-test. I The mRNA expression levels of Notch target genes were evaluated in SKOV3 cells treated with different concentrations of COMP (20 and 50 μg/ml) or BSA (50 μg/ml) as a control by RT-qPCR. The p-value was calculated by two-way ANOVA followed by Dunnett’s post-test. ACTB was used as a reference gene, and the relative mRNA expression was calculated using the 2^−ΔΔCt method. The data are representative of at least three independent experiments and graphs depict the mean with the standard deviation. P < 0.05 was considered statistically significant. (*, **, ***, ****, and ns indicate p < 0.05, p < 0.01, p < 0.001, p < 0.0001, and non-significant, respectively; NICD: Notch intracellular domain, NEXT: Notch extracellular truncated domain, FL: Full length)