Fig. 1: AKT/STAT3-induced VAL correlates with poor prognosis of LAD.

a Volcano plot shows significantly upregulated (red) and down-regulated (blue) lncRNAs in A549-myr-AKT1 cells as compared to A549-vector cells based on RNA sequencing. b The effect of overexpressing myr-AKT1 or distinct AKT inhibitors on VAL expression. c Schematic diagram of predicted binding sites for STAT3, p65, and HIF1α in the promoter/enhancer region of the VAL (LINC01546) gene. The predicted binding site containing wild-type or mutated binding motif for STAT3 is shown. d The effect of treatment of inhibitors for p65 (JSH-23), STAT3 (S3I-201), HIF1α (KC7F2), or mTOR (Rapamycin) on VAL expression. e, f Beas2B cells were transduced with the pLVX-TRE3G-myr-AKT1 Tet-on inducible system and stimulated with 2 μg/ml doxycycline (Dox) for 48 h, followed by treatment of AKT or STAT3 inhibitor. Expression of AKT1 and VAL in the indicated cells is shown. g, h The effect of myr-AKT1 overexpression followed by p65 inhibitor treatment on IL6 expression and secretion. i ChIP assays show the binding of STAT3 to the predicted promoter/enhancer region of the VAL gene in response to the indicated treatments. j Relative luciferase activities of the reporter constructs spanning wild-type or mutant predicted putative binding site for STAT3. k, l Analysis of VAL expression in 38 pairs of human LAD tissue (Tumor) and adjacent non-tumor tissue (Normal), and 46 cases of lung tissue and 495 cases of LAD tissue in the TCGA LAD datasets. m, n The expression levels of VAL in 34 pairs of LAD tissue and adjacent non-tumorous tissue using RNA in situ hybridization and representative images of VAL staining are shown. Scale bar: 100 μm. o, p Kaplan–Meier analysis (Log-rank test) of the 5-year overall survival (left panel) and recurrence-free survival (right panel) of LAD patients in the TCGA LAD datasets, who were divided into low or high VAL expression subgroups. Error bars represent the means ± SD derived from three independent experiments. Statistical analyses were performed by two-way ANOVA multiple comparison analysis (f–j), two-tailed unpaired Student’s t test (b, d, l), and two-tailed paired Student’s t test (k, n). n.s., not significant. Source data are provided as a Source Data file.