FIGURE 4.

Overexpression of MDA-9 activates ILK and NF-κB pathways. (A) Heatmap of top upregulated and downregulated genes in naive Alb/MDA-9 hepatocytes versus WT identified by RNA-seq. Spp1 is indicated by an asterix. (B) Signaling pathways significantly activated in naïve Alb/MDA-9 hepatocytes versus WT identified by IPA of RNA-seq data. Signaling pathways with −log(p value) of 2.5 and higher and z score of 2.0 and higher are shown. (C) Representative Western blot of the indicated proteins in WT and Alb/MDA-9 hepatocytes. β-Actin was used as loading control. (D) NF-κB luciferase reporter activity measured in WT and Alb/MDA-9 hepatocytes. Firefly luciferase activity was normalized by Renilla luciferase activity. Data represent mean ± SEM. *p < 0.01. E. Expression analysis of the indicated mRNAs by Q-RT-PCR in WT and Alb/MDA-9 hepatocytes. Data represent mean ± SEM. *p < 0.01. Alb, Albumin; A.U., arbitrary units; AKT, serine threonine kinase AKT; ERK, extracellular regulated kinase; FAT10, ubiquitin D; HA, hemagglutinin; ILK, integrin-linked kinase; iNOS, inducible nitric oxide synthase; IPA, ingenuity pathway analysis; MDA-9, Melanoma differentiation associated gene-9; MSP-RON, macrophage-stimulating protein-recepteur d’origine nantais; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; NO, nitric oxide; pGL3, empty pGL3-basic vector; PI3K, phosphatidylinositol 3-kinase; R.L.U., relative luciferase units; Q-RT-PCR, quantitative real time polymerase chain reaction; Rac, Rac family small GTPase; RNA-seq, RNA sequencing; ROS, reactive oxygen species; Spp1, Secreted phosphoprotein 1; TG, transgenic; TNFR1, TNF receptor 1; WT, wild-type.