TIGAR inhibits glucose-metabolism and cisplatin-chemosensitivity in human lung cancer cells

Abstract

TIGAR is an important factor associated with tumor glucose metabolism, but its function and underlying mechanism in human lung cancer remains unclear. Here, we analyzed the expression changes, prognosis, genetic alteration, related gene networks and metabolic pathways of TIGAR in lung cancer. The findings revealed that TIAGR level was augmented in LUAD and LUSC in comparison to the normal lung tissue. In addition, high TIAGR level was related to poorer outcome of patients with LUAD. Different alterations in TIGAR gene at various sites were observed in both LUAD and LUSC. The GO/KEGG analyses indicated that TIGAR affects the occurrence and progress of lung cancer through multiple metabolic pathways. Further, we established lung cancer cell models with TIGAR knockdown or overexpression to explore its effects on glucose metabolism, apoptosis and chemosensitivity. Our results indicated that TIGAR markedly inhibited glucose metabolism, ROS production, and susceptibility of lung cancer cells to cisplatin. Together, TIGAR plays a cancer-promoting role in lung cancer, which becomes a promising prognostic and therapeutic biomarker.

Keywords: Bioinformatics; Chemosensitivity; Lung cancer; Metabolism; TIGAR.

Fig. 1

TIGAR expression in lung cancer and its relationship with prognosis. A The levels of TIGAR mRNA in lung cancer (***: p < 0.001). B The levels of TIGAR mRNA in LUAD. (C) The levels of TIGAR mRNA in LUSC. D Protein expression levels of TIGAR in normal lung tissues (top), LUAD (middle) and LUSC (bottom) (IHC). E–G Association between TIGAR expression and OS in patients with lung cancer E, LUAD F and LUSC G

Fig. 2

Mutation feature of TIGAR in LUAD and LUSC. A Analysis of TIGAR alteration frequency and the mutation type in LUAD and LUSC. B Association between mRNA expression of TIGAR and its copy number in the LUAD and LUSC. C The mutation site of TIGAR in LUAD and LUSC. D Corresponding mutation site is shown in the 3D structure of TIGAR

Fig. 3

TIGAR -related gene enrichment and pathway analysis. A Protein-protein interaction network analysis of TIGAR. B–C KEGG B or GO C pathway analysis according to the TIGAR-binding and interacted genes in lung cancer. BP Biological Process, CC Cellular Component, MF Molecular Function

Fig. 4

TIGAR regulates glucose metabolism and affects the expression of glucose metabolism-related enzymes. The cells that infected by retrovirus containing TIGAR-shRNA or TIGAR-flag sequences were selected, and the glucose consumption of the cells were measured A, C; and the mRNA level of the indicated genes were tested by RT-PCR assay B, D. Graph bars represent the mean relative level of glucose consumption or mRNA from 3 independent experiments. Error bars indicate standard deviation. Student’s t-test p-value was considered to be statistically significant when it was < 0.05.The pLKO.1-shRNA and pBABE-puro vector systems were employed to construct plasmids for specific TIGAR knockdown and overexpression respectively. NC pLKO.1 negative control plasmid, pBABE pBABE-puro empty vector

Fig. 5

TIGAR regulates the level of ROS and apoptosis. A–B The level of ROS in TIGAR knockdown A or over-expression B cells were detected with DCFH-DA. C–D The apoptosis rate of the cells in TIGAR knockdown C or over-expression D were detected by flow cytometry. Graph bars represent the mean relative level of ROS or apoptosis rate from 3 independent experiments. Error bars indicate standard deviation. Student’s t-test p-value was statistically significant when it was < 0.05

Fig. 6

The responses of lung cancer cells to cisplatin. The A549 and H1299 cells were treated with 0, 10, 20 µM cisplatin for 24 h, the mRNA levels of the indicated genes were tested by qRT-PCR A, C, the protein levels of TIGAR were tested by Western Blot B, the glucose consumption D and the level of ROS E were also tested. The original blots of B are presented in Supplementary information file

Fig. 7

TIGAR reduces chemosensitivity of lung cancer cells to cisplatin. A-B The glucose consumption of TIGAR knockdown A or over-expression B cells treated with 20 µM cisplatin were detected. C–D The level of ROS in TIGAR knockdown C or over-expression D cells treated with 20 µM cisplatin were detected with DCFH-DA. E-F The TIGAR knockdown E or over-expressed F cells were treated with the indicated concentrations of cisplatin for 48 h, the survival of the cells was detected, and IC50 was calculated