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. 2016 Mar 8;7(15):20561–20573. doi: 10.18632/oncotarget.7987

Figure 1. RalA GTPase activity is increased in CML cells.

Figure 1

(A) RalA GTPase activity was determined by G-LISA™ in CML cell lines and three CML peripheral blood samples, compared to normal healthy blood controls. (B) RalA GTPase activity measured by G-LISA™ in K562 cells with overexpression of miR-181a or RalA siRNA compared to scrambled controls (SCR). (C) Confocal microscopy analysis of the Ral A localization in K562 cells. (D) BaF3 cells transfected with RalA plasmid, empty vector or blank were treated with different concentrations of Ara-C (1.2–2.8 μM) for 48 h and the viability of cells was determined by MTT assay. The preliminary experiment showed that overexpression of RalA could counteract Ara-C-induced cytotoxicity, and increase colony formation capacity in BaF3 cells. (E and F) 1000 BaF3 cells with RalA vector were mixed with RPMI-1640 medium containing 0.9% methylcellulose solution and 20% FBS, and seeded onto 24-well plates. Colony numbers were counted after 1 week. Histogram (E) shows the relative number of colonies per 1000 plated cells. Overexpression of RalA increases the colony-forming activity in BaF3 cells. (G) Xenograft model of CML with the human cell line K562 with RalA or empty vector in Balb/c mouse. 500,000 K562 cells transfected with RalA or empty vector were injected via intravenous tail vein injection into sublethally irradiated Balb/c recipient mice. Overall survival is shown by Kaplan–Meier analysis.