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CSIG-02. FOSL1 PROMOTES GLIOMA STEM CELLS TRAITS THROUGH INTERACTING WITH ITS BINDING PROTEIN PHOSPHORYLATED STAT3


 
 
 Glioma stem cells (GSCs) have key roles in tumorigenesis, progression, and resistance to conventional treatments for glioblastoma. We reported that TRPM7 channels, which fused with C-terminal kinase, endow glioma cells with stem cell-like capacities of self-renewal. Here, we investigated the role of TRPM7 downstream target gene FOSL1’s post transcription in glioma stemness.
 
 
 
 1) The immunohistochemical staining for FOSL1 was performed using the rabbit polyclonal anti-FOSL1 antibody in glioma tissue arrays from 76 Chinese patients and 10 normal individuals. The staining intensity of cells in TMA was evaluated in three different fields (≥100 cells/field). The semi-quantitative HSCORE was calculated for FOSL1. 2) Upon suppression of FOSL1 by siRNA FOSL1, GSC markers, CD133 expression was evaluated by flow cytometry using PE anti-human CD133 antibody, while ALDH1 enzymatic activity was assessed using an Aldefluor kit, in A172 and U87MG glioma cell lines and patient-derived xenoline PDX-L14 of proneural subtype. 3) To assay FOSL1 gene promoter activity, the 5’-flanking region of the human FOSL1 gene (1201bp) was inserted into the firefly luciferase reporter vector pGL3-Basic between MluI and BglII sites. For mutant construct, STAT3 binding sites were mutated using Q5 site-directed mutagenesis kit.
 
 
 
 1) Quantification of the IHC results showed that FOSL1’s positive nuclear staining was significantly higher in GBM (n=13, p=0.0005), grade III (n=12, p=0.0116), and grade II astrocytoma (n=51, p=0.0485) compared to that of normal brain tissue (n=10). 2) GSC markers CD133 and ALDH1 were positively associated with FOSL1 in glioma. 3) Phosphorylated STAT3 directly binds to the FOSL1 promoter and the activation levels of STAT3 have a positive effect on the transcriptional activity of FOSL1.
 
 
 
 Our results demonstrated the role of TRPM7-regulated STAT3 activation directly regulated FOSL1 gene transcription, which shed new light on TRPM7/FOSL1-directed therapeutics in glioma. (This study was supported by NIH NIGMS GM121230)
