Micro–adhesion rings surrounding TCR microclusters are essential for T cell activation

Files in this Data Supplement:

• Supplemental Materials (PDF)
• Video 1 -
  TCR-MCs transiently bordered by Pyk2 in the initial phase of activation. TIRF videos of TCR (H57)–Dy549 and Pyk2-GFP in AND-tg T cells placed on 1 µM or 10 nM Ag–loaded planer bilayer were taken at the rate of one frame per 5 s for 3 min. In the merged images, green and red show Pyk2-GFP and TCR, respectively. The fields of 18-µm squares for each color or merged videos are arranged.
• Video 2 -
  Different behaviors of ZAP70 and SLP76 clusters upon weak stimulation. TIRF videos of TCR (H57)–Dy549, ZAP70-GFP, SLP76-GFP, and LAT-GFP in T cells placed on 1 µM or 10 nM Ag–loaded planer bilayer were taken at the rate of one frame per 5 s for 6 min. The fields of 18-µm squares for each color or merged videos are arranged.
• Video 3 -
  F-actin transiently accumulated at TCR-MCs in the initial phase of activation. TIRF videos of TCR (H57)–Dy549 and Lifeact-GFP in T cells placed on 1 µM Ag–loaded planer bilayer were taken at the rate of one frame per 5 s for 5 min. In the merged images, green and red show Lifeact-GFP and TCRs, respectively. The fields of 18-µm squares for each color or merged videos are arranged.
• Video 4 -
  Disappearance of SLP76 clusters and remaining TCR-MCs and ZAP70 clusters by CytD treatment. TIRF videos of TCR (H57)–Dy549, ZAP70-GFP, and SLP76-GFP in T cells placed on the planar bilayer containing 1 µM Ag taken at the rate of one frame per 5 s for 5 min. At the 11th frame, CytD was added at a final concentration of 10 µM. The fields of 14.5-µm squares are arranged.
• Video 5 -
  The unstable contacts and impaired IS formation of dKD cells on the planer bilayer. TIRF videos of TCR (H57)–Dy549 in shCtrl and dKD cells on 1 µM or 10 nM Ag–loaded planer bilayer were taken at the rate of one frame per 5 s for 5 min. The fields of 22-µm squares are arranged.
• Video 6 -
  The unstable contacts and impaired IS formation of dKD cells upon interaction with APCs. The videos of TCR (H57)–Dy549 and a differential interference contrast image in shCtrl and dKD cells with 1 µM Ag–loaded activated B cells were taken at the rate of one frame per 10 s for 8.5 min. The fields of 14-µm squares are arranged.