TPC2 mediates new mechanisms of platelet dense granule membrane dynamics through regulation of Ca2+ release

Supplemental Materials

This article contains the following supporting material:

  • Supplemental Materials
  • video 7 - Video 7. Tubule connections mediate transfer of membrane proteins between PDGs. Original movie Figure 7E. MEG‐01 cells were transfected with LAMP2‐PAGFP and TPC2‐Cherry. The video shows the transferring of fluorescent LAMP2‐PAGFP molecules along a tubule connecting a irradiated PDG (at time 0 sec) with a non‐photoactivated granule. Time is seconds.
  • Video 3 - Video 3. TPC2 is involved in PDG membrane fusion. Original movie Figure 6A. Confocal fluorescent microscopy image of a representative MEG‐01 cell expressing wt TPC2‐GCaMP6 and VMAT2‐Cherry. Time is min:sec.
  • video 4 - Video 4. TPC2 is involved in PDG membrane fusion. Original movie insets Figure 6B. PDGs in a MEG‐01 cell expressing TPC2‐cherry were loaded with the fluid phase marker Dextran Alexa488. Time is min:sec.
  • video 6 - Video 6. “Kiss‐and‐run” mediates transfer of membrane proteins between PDGs. Original movie Figure 7C. MEG‐01 cells were transfected with LAMP2‐PAGFP and VMAT2‐Cherry. A single PDG was photoactivated and the video shows transfer of fluorescent LAMP2‐PAGFP into a non‐photoactivated granule. Time is seconds.
  • video 5 - Video 5. TPC2 is involved in PDG membrane fusion. Original movie Figure S9. Primary MKs were transduced with TPC2‐Cherry and PDGs in these cells were labeled with the DG‐specific dye mepacrine. Time is min:sec.
  • Video 1 - Video 1. TPC2‐GCaMP6 can be used to monitor changes in [Ca2+]i. The ability of TPC2‐GCaMP6 to detect changes in [Ca2+]i in real time was tested in MEG‐01 cells stimulated with Thrombin and Thapsigargin using a microfluidic system. Similar to synthetic calcium indicators such as Fura2‐AM the images show TPC2‐GCaMP6 is able to sense the variations in [Ca2+]i triggered by these reagents. Time is min:sec.
  • Video 2 - Video 2. TPC2 is involved in the formation of perigranular Ca2+ nanodomains. Original movie Figure 5C. Confocal fluorescent microscopy images at different time points of MEG‐01 cells co‐expressing TPC2‐GCaMP6 and VMAT2‐Cherry. Time is min:sec.