Vertebrate MIM loss of function results in increased endocytosis and reduced directional migration. (A and B) Quantification of the ratio of internalized to surface-bound Transferrin (A) of EGF (B) in ptc^−/− MEFs treated with indicated siRNAs. Data are represented as the mean ± SEM from three separate experiments (*, P < 0.01; t test). (C and D) Quantification of the amount of recycled Transferrin (C) or EGF (D) in the medium of ptc^−/− MEFs treated with indicated siRNAs. Data are represented as the mean ± SEM from three separate experiments. Inset in C shows immunoblots of protein level knockdown after treatment with indicated siRNAs. (E) MIM knockdown alters EGF-induced directional cell migration. Transwell migration assays showing alterations in the migration of ptc^−/− MEFs through a permeable membrane in response to an EGF or PDGF gradient over the course of 8 h. Data are represented as the mean ± SEM from three separate experiments (*, P < 0.01; t test). (F) EGFR levels in siRNA-treated cells show that si-MIM–treated cells do not display enhanced degradation of the EGF receptor over time. (G) Phospho-ERK1 and 2 levels in cells treated with control or MIM siRNA. MIM knockdown results in prolonged levels of pERK1/2 after the addition of EGF to the cell medium. (H and I) Quantification of immunoblots used for F and G. Data are represented as the mean ± SEM from three separate experiments (*, P < 0.01; t test).