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. 2015 May 5;35(11):1898–1914. doi: 10.1128/MCB.01398-14

FIG 3.

FIG 3

H-Ras–GDP and H-Ras–GTP occupy different PM microdomains. (A) Endogenous H-Ras localization at DRMs and SFs in serum-starved or EGF-stimulated (st.; 100 ng/ml, 10 min) HeLa cells. (B) As described in the legend to panel A, the distribution of endogenous H-Ras and K-Ras was analyzed in HCT116 cells, including proliferating cells (prolif.). (C and D) Effects of EGF on H-Ras sublocalization in HeLa (C) and HCT116 (D) cells analyzed by FRET. H-Ras–Cerulean was tested against the probes specific for DRMs (LCK-Venus [LCK-v]) and SFs (Venus-CD8 [CD8-v]). (E) Controls for the site-specific FRET probes CD8 and LCK. Cells transfected with control plasmids show minimum (CTV) and maximum (C5V) FRET efficiencies. Results show the mean ± SEM from at least five experiments. **, P < 0.01 with 95% confidence intervals; ***, P < 0.001 with 95% confidence intervals. LCKc, LCK-Cerulean. (F) Endogenous H-Ras distribution in MCA3D and HEK293T cells stably expressing the exchange factors SOS1 (SOS) and RasGRF1 (GRF). Cells were serum starved overnight before processing.