Supplement to Tiam–Rac signaling mediates trans-endocytosis of ephrin receptor EphB2 and is important for cell repulsion

Files in this Data Supplement:

Supplemental Materials (PDF)
Video 1 -
F-actin is enriched at sites of EphB2 trans-endocytosis into SKN cells. Representative movie from Fig. 1 (C and E–G) showing EphB2ΔC-GFP (green) trans-endocytosis into ephrinB⁺ SKN cell overexpressing LifeAct-mCherry (red). Cells were co-cultured and imaged live once contact occurred. EphB2ΔC-GFP–positive vesicles (green and yellow puncta) can be seen in the SKN cell. Time stamp shows hours:minutes:seconds.
Video 2 -
Rac inhibition blocks F-actin enrichment at sites of EphB2 contact with SKN cells. Representative movie from Fig. 1 (D, H, and I) and Fig. S1 B showing EphB2ΔC-GFP (green) trans-endocytosis into ephrinB⁺ SKN cell overexpressing LifeAct-mCherry (red) exposed to 10 µM EHT1864. Cells were co-cultured and imaged live once contact occurred. Time stamp shows hours:minutes:seconds.
Video 3 -
EphB2ΔC trans-endocytosis into primary neurons. Representative movie for Fig. 3 (A and B) showing EphB2ΔC trans-endocytosis into embryonic day 15.5 mouse cortical neurons in vehicle control condition. Neurons were stained with CellTracker green, co-cultured with EphB2ΔC-mCherry–positive HeLa cells, and imaged live. Time stamp shows hours:minutes:seconds.
Video 4 -
Rac inhibition blocks EphB2ΔC trans-endocytosis into primary neurons. Representative movie for Fig. 3 (A and B) showing prolonged contact between EphB2ΔC⁺ cells and embryonic day 15.5 mouse cortical neurons after 10 µM EHT1864 treatment. Neurons were stained with CellTracker green, co-cultured with EphB2ΔC-mCherry–positive HeLa cells, and imaged live. Time stamp shows hours:minutes:seconds.
Video 5 -
EphB2ΔC-triggered cell repulsion in SKN cells. Representative movie for Fig. 8 (A and B) showing EphB2-triggered cell repulsion in ephrinB⁺ SKN cells in the vehicle control condition. SKN cells were transfected with mCherry, co-cultured with EphB2ΔC-GFP–positive HeLa cells, and imaged live. Time stamp shows hours:minutes. Bar, 20 µm.
Video 6 -
Rac inhibition blocks EphB2ΔC-triggered cell repulsion. Representative movie for Fig. 8 (A and B) showing prolonged contact between EphB2⁺ cells and ephrinB⁺ SKN cells after 10-µM EHT1864 treatment. SKN cells were transfected with mCherry, co-cultured with EphB2ΔC-GFP–positive HeLa cells, and imaged live. Time stamp shows hours:minutes. Bar, 20 µm.
Table S1 (Excel file) -
Results from the Rho GEF and GAP screen and the entire data set for the siRNA screen from Figs. 6 and S4. Included in the table are the siRNA sequences for each gene, the average z-score from the two runs, and the combined gene average. GTPase specificity is also labeled.