Fig. 4: AHR regulates ASNS expression in an ATF4-dependent manner and affects MMP expression and activity.
From: Functional screening identifies aryl hydrocarbon receptor as suppressor of lung cancer metastasis
Target gene expression of three independent clones of H1975 expressing shAHR (K1-K3) and H1975 shScr control cells (shScr) was analyzed after treatment with omeprazole (omep, 200 µM, 48 h) or DMSO (−) by qRT-PCR (a, b) and immunoblotting (c–e). a qRT-PCR analysis of ASNS mRNA expression. b qRT-PCR analysis of MMP24 mRNA expression. c–e Immunoblotting analysis of AHR, ASNS (c), MMP24 (d), and ATF4 (e) protein expression. f Prior to treatment with omeprazole (omep, 200 µM, 48 h), H1975, A549, and H1299 were transfected with siRNAs targeting ATF4 or a non-targeting control (siCtrl). Expression of ASNS and ATF4 was assessed using immunoblotting. g qRT-PCR analysis of ASNS and ATF4 mRNA levels relative to siCtrl control. Target gene expression was normalized to two (ACTB, GAPDH) or three (ACTB, GAPDH, HPRT1) housekeeping genes (HKG) as indicated. h Gelatine zymography of MMP2 and MMP9 activity. i Semiquantitative analysis of gelatinolytic bands from MMP9 (ImageJ). Data are shown as mean ± SD. Significance was assessed using one-way ANOVA.
