Figure 6.
GIT1 is an SRC effector protein that regulates YAP/TAZ activity. A–C, the indicated cell lines were co-transfected with either control empty vector or SRCY527F and GIT1 or Cool-1 DNA and then assayed for YAP/TAZ-TEAD transcriptional activity 24 h later. D and E, A375 or A375-pTRIPZ-SRCY527F cells were transfected with FLAG-tagged GIT1 or FLAG-tagged Cool-1. After 24 h, A375 cells were treated with either DMSO (−) or 500 nm dasatinib (+) for 1 h (left), and A375-pTRIPZ-SRCY527F cells were treated with water (−) or 0.5 μg/ml doxycycline (+) for 12 h (right). Western blots were then performed on whole-cell lysate (WCL) or following IP with an anti-FLAG antibody for phosphorylated tyrosine, GIT1, Cool-1 pSFK, or GAPDH. As a control, the indicated sample was also incubated with calf intestinal phosphatase (CIP) prior to loading on the gel. A, n = 4 (one experiment performed in quadruplicate). Scatter plots show mean ± S.D. (error bars), where each dot is an independent experiment. Statistical significance was tested using Student's t test with Bonferroni correction for multiple comparisons. *, p ≤ 0.05; **, p ≤ 0.01; ***, p ≤ 0.001; ****, p ≤ 0.0001.