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Significance How endocytosis regulates intracellular signaling is a major unsolved question. In this study, we labeled Ras, which plays a central role in normal and oncogenic signaling, with the fluorescent protein by gene editing, and for the first time (to our knowledge) examined the localization of endogenous Ras in living cells stimulated with epidermal growth factor (EGF). Microscopy imaging of living cells demonstrated that, although activated EGF receptors are rapidly internalized into endosomes, Ras is not present in these endosomes and mainly located in the plasma membrane. Therefore, EGF receptors signal to MAP kinases through Ras exclusively from the plasma membrane.

Endocytosis separates EGF receptors from endogenous fluorescently labeled HRas and diminishes receptor signaling to MAP kinases in endosomes