Figure 2
From: Optogenetically controlled RAF to characterize BRAF and CRAF protein kinase inhibitors
OptoRAF systems activate MAPK pathway (A) HEK293T cells were transfected with the plasmids as indicated and incubated for about 50 h in darkness (−) or illuminated for 5 min with light of 460 nm (+) before lysis of the cells. Cell lysates were immunoblotted with the respective antibodies to detect CRAF and BRAF and to analyze MEK and ERK activation by phospho-specific antibodies (pMEK1/2 and pERK1/2, respectively). β-actin was used as loading control. For quantification of the pERK1/2 levels see Supplementary Fig. S3. (B) HEK293T cells expressing the RAF constructs as indicated were incubated for 24 h and subsequently incubated for further 24 h in darkness (black columns) or exposed to continuous blue light (blue columns). SEAP activity was determined from the supernatants collected.
