Fig. 4: MEK2-R67 methylation is its functional active site.

A Plate cloning test to examine how ME2 mutation and wild type affect the capacity of HCC cells to form colonies after ME2 knockdown. B The CCK-8 experiment to examine how ME2 wild-type and mutation affect the activity of HCC cells after ME2 knockdown. C Cell suspension culture analysis of how ME2 wild type and mutation affect the stemness of ME2-knocked down HCC cells. D Transwell Migration and Invasion Assay to examine how ME2 wild type and mutation affect invasion and metastasis of HCC cells that have had ME2 knocked down. E Cell scratch experiment to examine how the wild type and mutant forms of ME2 affect the capacity of HCC cells to migrate. F–H. The effects of ME2 wild type and mutant on the tumorigenicity of HCC cells with ME2 knockdown were examined using a subcutaneous tumorigenicity test. Thymus-free nude mice (n = 5/group) received subcutaneous injections of HepG2 and Hep3B cells (2 × 10⁶) with varying ME2 status. After 28 days of injection, the mice were sacrificed, and the tumor development was analyzed (F). Tumor volume (H) and weight (G) were assessed. I, J. To assess the expression and statistical quantification of KI67 and PCNA in the xenograft tumor tissues of each group, immunohistochemical procedures were performed. K GSH/GSSG and NADPH/NADP+ ratios in every set of xenograft tumor tissues.