Fig. 4.

SLC2A3 (glucose transporter 3 (GLUT3)) is a downstream gene in temozolomide (TMZ)-induced DNA damage-inducible transcript 4 (DDIT4) signaling. (A) Effects of DDIT4 overexpression on mRNA levels of 5 DDIT4-associated genes. (B) Only GLUT3 upregulation in microarray data of TMZ-treated cells. (C) TMZ treatment increased GLUT3 mRNA levels in a dose-dependent manner. (D) TMZ treatment and overexpression of DDIT4 enhanced GLUT3 protein levels. (E) Overexpression and knockdown of DDIT4 influenced TMZ-induced GLUT3 expression. (F) Knockdown of GLUT3 enhanced TMZ-induced caspase-3 activation. Overexpression of GLUT3 attenuated the effects of DDIT4 depletion of enhancing TMZ cytotoxicity in U87-MG (G) and T98G cells (H). After transfection with the indicated dose or 2 μg of DDIT4 plasmids for 24 h, indicated dose or 200 μM TMZ was added for another 72 h. Relative mRNA and protein levels, and cell viability were respectively measured by real-time PCR, immunoblot, and MTT assays. Caspase-3 activity was measured by using commercial kit according to the manufacturer’s instructions listed in the “Materials and Methods” section. Data are the mean ± SD of 3 experiments. *p < 0.05. A significant positive correlation existed between DDIT4 and GLUT3 in glioblastoma multiforme (GBM) tissue arrays (I) and The Cancer Genome Atlas (TCGA) pan-cancer analyses (J). BLCA = urothelial bladder carcinoma; BRCA = breast invasive carcinoma; COAD = colon adenocarcinoma; HNSC = head-neck squamous cell carcinoma; KIRC = kidney renal clear cell carcinoma; LUAD = lung adenocarcinoma; LUSC = lung squamous cell carcinoma; OV = ovarian cancer; UCEC = uterine corpus endometrial carcinoma