Figure 4. Silencing of MYSM1 in A375 and SK-MEL-28 melanoma cells affects tumor cell survival and proliferation.
The stable cell lines designated as clones d, e, and h are mixed populations derived from two different parental melanoma cell lines, A375 (black bars) and SK-MEL-28 (grey bars), lentivirally transduced to express MYSM1 shRNA clones a-h and GFP. Scrambled control A375 and SK-MEL-28 cells were stably transduced with scrambled control RNA. Bar graphs and standard deviations in each panel represent results of three independent experiments. (A) Knockdown efficiency of MYSM1 shRNAs in sorted GFP+ A375 and SK-MEL-28 melanoma cells compared with scrambled control RNA and parental cells analyzed by FACS and qPCR, respectively. (B) MTT assay of melanoma cell viability upon MYSM1 knockdown compared with expression of scrambled control RNA. (C) Annexin V-apoptosis assay of GFP-positive melanoma cells after MYSM1 knockdown compared with scrambled control cells by FACS-analysis. (D) Softagar-assay of MYSM1-silenced in comparison with parental and scrambled control RNA-expressing A375 and SK-MEL-28 melanoma cells. In each case, 103 cells/dish were seeded and colonies with diameter > 125μm were counted after 14 days of incubation.