Figure 7.
Figure 7. Myo1b is required to form filopodia at EphB2–ephrinB1 cell interface. (A) YFP-EphB2-Hek293T transfected with control siRNA cells were cultivated with Cherry-ephrinB1-Hek293T cells or themselves in Ibidi inserts and their behavior was monitored by time-lapse microscopy (Video 6). Representative frames for YFP-EphB2-Hek293T cells cultivated with other YFP-EphB2-Hek293T cells or with Cherry-ephrinB1-Hek293T cells are shown. Bars, 10 µm. (B) Distribution of the number of filopodia observed per frame normalized to the total number of frames analyzed (1 frame/5 min during 350 min) and distribution of the size of filopodia observed when YFP-EphB2-Hek293T cells transfected with control siRNA were facing themselves (11 videos) or Cherry-ephrinB1-Hek293T cells (11 videos). The number of frames with more than four filopodia and the length of the filopodia increase when YFP-EphB2-Hek293T cells contact ephrinB1-Hek293T cells. (C) Distribution of the number of filopodia observed per frame normalized to the total number of frames analyzed (1 frame/5 min during 350 min) and distribution of the size of filopodia observed in YFP-EphB2-Hek293T cells treated with control (11 videos) or Myo1b (12 videos) siRNAs and facing Cherry-ephrinB1-Hek293T cells. Myo1b KD increases the number of frames without filopodia but does not affect the size of the remaining filopodia. (D) YFP-EphB2-Hek293T cells treated with control or Myo1b siRNA were cultivated with Cherry-ephrinB1-Hek293T cells in Ibidi inserts and their behavior was monitored by time-lapse microscopy (Videos 7 and 8). Phase contrast of the first image and sequence of YFP fluorescent images illustrating the formation of filopodia after the first contact via lamellipodia are shown in the presence or absence of Myo1b. Arrows mark the elongating filopodia. Bar, 10 µm. (E) Time observed between the first contact formed by YFP-EphB2-Hek293T cells treated with control or Myo1b siRNA and Cherry-ephrinB1-Hek293T cells and the observation of the first filopodia was quantified and presented as box plots. Mann-Whitney test = 0.04.

Myo1b is required to form filopodia at EphB2–ephrinB1 cell interface. (A) YFP-EphB2-Hek293T transfected with control siRNA cells were cultivated with Cherry-ephrinB1-Hek293T cells or themselves in Ibidi inserts and their behavior was monitored by time-lapse microscopy (Video 6). Representative frames for YFP-EphB2-Hek293T cells cultivated with other YFP-EphB2-Hek293T cells or with Cherry-ephrinB1-Hek293T cells are shown. Bars, 10 µm. (B) Distribution of the number of filopodia observed per frame normalized to the total number of frames analyzed (1 frame/5 min during 350 min) and distribution of the size of filopodia observed when YFP-EphB2-Hek293T cells transfected with control siRNA were facing themselves (11 videos) or Cherry-ephrinB1-Hek293T cells (11 videos). The number of frames with more than four filopodia and the length of the filopodia increase when YFP-EphB2-Hek293T cells contact ephrinB1-Hek293T cells. (C) Distribution of the number of filopodia observed per frame normalized to the total number of frames analyzed (1 frame/5 min during 350 min) and distribution of the size of filopodia observed in YFP-EphB2-Hek293T cells treated with control (11 videos) or Myo1b (12 videos) siRNAs and facing Cherry-ephrinB1-Hek293T cells. Myo1b KD increases the number of frames without filopodia but does not affect the size of the remaining filopodia. (D) YFP-EphB2-Hek293T cells treated with control or Myo1b siRNA were cultivated with Cherry-ephrinB1-Hek293T cells in Ibidi inserts and their behavior was monitored by time-lapse microscopy (Videos 7 and 8). Phase contrast of the first image and sequence of YFP fluorescent images illustrating the formation of filopodia after the first contact via lamellipodia are shown in the presence or absence of Myo1b. Arrows mark the elongating filopodia. Bar, 10 µm. (E) Time observed between the first contact formed by YFP-EphB2-Hek293T cells treated with control or Myo1b siRNA and Cherry-ephrinB1-Hek293T cells and the observation of the first filopodia was quantified and presented as box plots. Mann-Whitney test = 0.04.

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