Fig. 2: Restoring IRAK-M expression in human melanoma cell lines induces cell death.

a RAK-M protein level was determined by western blot in human melanocytes and melanoma cell lines transfected with empty vector or IRAK-M construct for 24 h. Blots are representative of at least two independent experiments. b Human melanocytes and melanoma cell lines were transfected with a plasmid control or pIRAK-M, and apoptosis was measured by flow cytometry (Annexin V staining) 24 hours later (n = 3 per group). Results are shown as mean ± SEM. **p < 0.01 by two-tailed Student’s t-test. Data are representative of at least three experiments, each yielding identical results. c Expression levels of apoptosis-related proteins in melanoma cells transfected for 24 h were evaluated by western blot. Blots shown are representative of three independent experiments. d The treatment effects of 20 µM Z-DEVD-FMK (a caspase-3 inhibitor), Z-IETD-FMK (a caspase-8 inhibitor), or Z-LEHD-FMK (a caspase-9 inhibitor) in IRAK-M transfected melanoma cells for 24 h were determined using flow cytometric analysis of apoptosis (n = 3 per group). Results are presented as mean ± SEM. *p < 0.05 by two-tailed Student’s t-test. Data presented are representative of two independent experiments. e, f Melanocytes and melanoma cell line RPMI7951 were transfected with psiRNA-Control or psiRNA-hIRAK-M vector for 24 h, followed by e flow cytometric analysis of apoptosis (n = 3 per group) and f western blot. Results are presented as mean ± SEM. Data shown are representative of two independent experiments.