Abstract
Background
Multi-drug resistance (MDR) remains a significant barrier to effective chemotherapy and results in poor prognosis of gastric cancer (GC) patients. Exploring the mechanism of MDR is of great significance for identifying biomarkers of resistance and developing new treatment strategies.
Methods
The combination of multi-omics and bio-informatics analysis with experimental validation was adopted to explore the mechanism of MDR in gastric cancer (GC). Multi-omics analysis includes transcriptome (RNA-Seq) and proteome (iTRAQ-MS and HLC-MS) analysis. Bio-informatics analysis includes a series of differential analysis, enrichment analysis, correlation analysis, survival analysis and molecular docking. Experimental validation includes quantitative real-time PCR (RT-qPCR), Western blot, immunofluorescence (IF), immunohistochemistry (IHC), multiplex immunohistochemistry (mIHC), CCK−8 assay, Clone formation experiment, Flow cytometry, Luciferase reporter assay, RNA stability assay, co-immunoprecipitation (Co-IP), RNA-binding protein immunoprecipitation (RIP), Chromatin immunoprecipitation (ChIP), RNA pull down assay and animal studies.
Results
In our study, we found that phosphoglycerate dehydrogenase (PHGDH), the key rate-limiting enzyme in the serine synthesis pathway, was significantly up-regulated in MDR GC cells. PHGDH, acting its non-canonical function, was found out to promote MDR by promoting the Wnt/β-catenin signaling pathway mediated by transcription factor 7 like 2 (TCF7L2), a pivotal transcription factor in the Wnt pathway. Specifically, PHGDH stabilized TCF7L2 mRNA by interacting with insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1), a key m6A reader. PHGDH/IGF2BP1 interaction was m6A-dependent and led to the enhanced TCF7L2 mRNA stability and thus its up-regulation. Moreover, TCF7L2 bound to the PHGDH promoter and regulated PHGDH expression, acting in a feedback way.
Conclusions
The PHGDH/IGF2BP1-TCF7L2 axis plays a vital role in the MDR of GC and correlates with poor prognosis.