The Effect And Mechanism Of Cytoskeletal Protein Coronin 3 On EMT Of Glioma T98G Cells

Objective:This study intends to investigate whether Coronin 3 is involved in the EMT process of GBM through in vitro cell experiments,and clarify its influence on cell polarity change and cell motor ability.Identify possible signaling pathways and factors associated with Coronin 3 in EMT.This study can clarify the function and mechanism of Coronin 3 in GBM,and also provide new targets and ideas for clinical treatment of glioma.Methods:Glioma T98G cells were cultured in vitro,the cells were divided into 4 groups,and the cells were transferred at the right time according to the groups.The 4 groups were wild-type T98G cell group(control),NC silencing group,Coronin 3-siRNA1 group and Coronin 3,siRNA2 group.Firstly,the knockdown effect was verified by Western blot and qRT-PCR.Then,the effect of knocking down Coronin 3 on cell migration was verified by wound healing assay.On this basis,we compared the expression of EMT-related proteins--epithelial cadherin,interstitial cadherin and vimentin in the above four groups of cells,so as to explore whether Coronin 3 is involved in EMT.At the same time,the effects of low expression of Coronin 3 on the polarity protein markers GM130,MUC1,cell polarity and the infection structure of cytoskeleton in glioblastoma T98G were further detected by cellular immunofluorescence method and phyllopeptides,so as to clarify the specific stage in which the actin skeleton remodeling function of Coronin 3 affects EMT.Finally,the TCGA database was used to download the GBM mRNA expression profile data,and Pearson correlation analysis was used to screen the EMT classical factors closely positively correlated with Coronin 3 through R software,and in vitro Western blot test and immunofluorescence test were used to verify this.Results:According to the analysis of wound healing assay,the migration ability of T98G cells was significantly inhibited after Coronin 3 expression was knocked down.Meanwhile,Western blot results showed that the expression of EMT-related epithelial cadherin(E-cadherin)was increased after Coronin 3 expression was lowered.The interstitial expression of N-cadherin and vimentin decreased.On this basis,we found that the expression level and position of polar protein markers GM130 and MUC1 did not change significantly after knocking down Coronin 3 by cell immunofluorescence assay.The results showed that the decreased expression of Coronin 3 inhibited the expression of cytoskeleton protein F-actin.Using R software analysis,we found that ZEB2 was closely positively correlated with Coronin 3,and Western blot and immunofluorescence experiments confirmed that the expression of ZEB2 decreased after the reduction of Coronin 3 expression.Conclusion:The results of this study confirmed that Coronin 3 knockdown can not only reduce the migration ability of T98G cells,but also inhibit the EMT of T98G cells in vitro,and this change also reduces the expression of cytoskeletal protein F-actin.Finally,we found that Coronin 3 was positively correlated with the classical EMT factor ZEB2,suggesting that Coronin 3 May be involved in the EMT process of glioma through ZEB2.This finding provides new ideas for the treatment of glioma,but further experiments are needed to understand the specific mechanism.