Figure 7.
EGFR and ligand-engaged EGFR in the basal PM detected virtually the same compartment sizes as those found with TfR and Cy3-DOPE, supporting the PM compartmentalization, and the dwell lifetime of the engaged EGFR was longer than that of non-engaged EGFR (same compartment sizes). Following the stimulation with 10 nM EGF, microscope observations were performed between 2.5 and 5 min after the EGF addition. (A) Typical ultrafast (6 kHz) single fluorescent-molecule trajectories of EGFR, before and after EGF stimulation (n = 41 and 39 trajectories, respectively). (B) Distributions of the compartment sizes detected by the single-molecule diffusion of EGFR in the basal PM, before (top) and after (bottom) EGF stimulation. Arrowheads indicate the median values of 106 nm for both before and after stimulation. (C) Distributions of the EGFR residency times within a compartment determined by the TILD analysis, with the best-fit exponential curves, providing the dwell lifetimes (τ). Before (top) and after (bottom) EGF stimulation. Statistically significant difference between before and after stimulation with P = 0.044, using the log-rank test. (D) Distributions of DMACRO for EGFR in the basal PM determined by the hop-diffusion fitting of the MSD-∆t plot for each EGFR molecule (shaded histogram, before stimulation; green open histogram, after stimulation). Arrowheads indicate the median values. DMACRO was reduced by a factor of 1.7 after stimulation.The statistical test methods, parameters (number of experiments), and P values are summarized in Table 3.

EGFR and ligand-engaged EGFR in the basal PM detected virtually the same compartment sizes as those found with TfR and Cy3-DOPE, supporting the PM compartmentalization, and the dwell lifetime of the engaged EGFR was longer than that of non-engaged EGFR (same compartment sizes). Following the stimulation with 10 nM EGF, microscope observations were performed between 2.5 and 5 min after the EGF addition. (A) Typical ultrafast (6 kHz) single fluorescent-molecule trajectories of EGFR, before and after EGF stimulation (n = 41 and 39 trajectories, respectively). (B) Distributions of the compartment sizes detected by the single-molecule diffusion of EGFR in the basal PM, before (top) and after (bottom) EGF stimulation. Arrowheads indicate the median values of 106 nm for both before and after stimulation. (C) Distributions of the EGFR residency times within a compartment determined by the TILD analysis, with the best-fit exponential curves, providing the dwell lifetimes (τ). Before (top) and after (bottom) EGF stimulation. Statistically significant difference between before and after stimulation with P = 0.044, using the log-rank test. (D) Distributions of DMACRO for EGFR in the basal PM determined by the hop-diffusion fitting of the MSD-∆t plot for each EGFR molecule (shaded histogram, before stimulation; green open histogram, after stimulation). Arrowheads indicate the median values. DMACRO was reduced by a factor of 1.7 after stimulation.The statistical test methods, parameters (number of experiments), and P values are summarized in Table 3.

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