Effect of PAK4-LIMK1-cofilin signaling on non-small-cell lung cancer mi-gration and invasion / 中国病理生理杂志

ABSTRACT

AIM:To explore the mechanism of p21-activated kinase 4 (PAK4) on non-small-cell lung cancer (NSCLC) migration and invasion.METHODS:After A549 and NCI-H520 cell lines were transfected with PAK4-siRNA or negative control , the expression of PAK4 at mRNA and protein levels was detected by real-time PCR and Western blot , re-spectively .The invasion and migration of A 549 cells and NCI-H520 cells were measured by Matrigel invasion assay and Transwell migration assay.LIMK1, cofilin, and their respective phosphorylation were examined by Western blot .The inter-action of PAK4 and LIMK1 was investigated by co-immunoprecipitation assay .The relationship between PAK4 and LIMK1 phosphorylation was examined by a protein kinase assay in the A 549 cells and NCI-H520 cells.The expression of PAK4 and p-LIMK1 in 10 human NSCLC tissues was examined by Western blot .A549 cells and NCI-H520 cells were individually or commonly transfected with PAK4-siRNA or LIMK1 plasmid in order to observe the cell migration and invasion .RE-SULTSAfter A549 cells and NCI-H520 cells were transfected with PAK4-siRNA for 48 h, the expression of PAK4 at mR-NA and protein levels , and the numbers of invasion and migration cells in PAK 4-siRNA group were lower than those in con-trol group.Compared with control group , the phosphorylation of LIMK1 and cofilin was lower in PAK4-siRNA group, whereas the total expression levels of LIMK 1 and cofilin did not change .The results of co-immunoprecipitation assays showed that PAK4 specifically interacted with LIMK1 in A549 and NCI-H520 cells.LIMK1 phosphorylation in the presence of PAK4 (K350M) was significantly lower than that in the presence of PAK 4 (WT) or PAK4 (S445N) in the protein ki-nase assay.The PAK4 upregulation was positively correlated with the level of p-LIMK1 (P<0.05).After A549 cells and NCI-H520 cells were co-transfected with PAK4-siRNA and LIMK1 plasmid, the migration and invasion cell numbers in co-transfection group were higher than those in PAK 4-siRNA transfection group .CONCLUSION: PAK4 promotes the inva-sive and migratory abilities of NSCLC , which is mediated by LIMK 1 phosphorylation .