Fig 2. Effect of methyl sulfone on hypoxia-dependent and -independent expression of proteins in metastatic cells.

66cl-4 breast cancer cells and M3 melanoma cells were grown under normoxic conditions in the presence and absence of methyl sulfone. When cells became confluent, half of the control plates and methyl sulfone-treated plates were placed under hypoxia. The remaining plates continued incubation under normoxic conditions. This led to four samples: control-normoxia (CN), control-hypoxia (CH), methyl sulfone-normoxia (MN), and methyl sulfone-hypoxia (MH). After six hours, cells were immediately put on ice, lysed, and prepared for immunoblot analysis. ImageJ determined relative levels within each set of the four samples. The most intense band was set at 100. Equal loading of protein was assessed by immunoblot analysis of YAP protein in 66cl-4 lysates and tubulin protein in M3 lysates. The immunoblots shown in Fig 2 were obtained from one lysate experiment for each cell line.