BLOC-2 interacts with the exocyst complex, which is essential for WPB biogenesis. (A) IPs of HUVEC lysates with anti-HPS6 (i) or EXOC4 (ii) antibodies resolved by SDS-PAGE and immunoblotted with HPS6 and EXOC4 antibodies. HUVEC lysate (∼5% of IP) used as input and IP with polyclonal IgG used as a control for both immunoblots. (B) IF analyses of EXOC4-depleted and control HUVECs labeled with anti-VWF antibody and counterstained with DAPI. As for BLOC-2–depleted cells, only rounded, immature WPBs, which are clumped perinuclearly are evident, highlighted by insets. Scale bars represent 10 μm. (C) Fifteen BLOC-2–depleted and control cells were randomly selected and total number of WPBs > 1.5 μM in length were determined for 2 independent studies (***P < .001). (D) EXOC4-depleted and control HUVECs dual labeled with anti-VWF and CD63 antibodies were analyzed by IF, counterstained with DAPI. The insets highlight colabeling of mature WPBs and CD63 in control cells, absent in EXOC4-depleted cells. Scale bars represent 10 μm. (E) GFP-CD63–expressing HUVECs analyzed by IF 48 hours after transfection with control or EXOC4 siRNA. Arrows show GFP-CD63 labeling of mature WPBs in control cells (i), absent in EXOC4-depleted cells (ii). Inset highlights mature WPBs. Scale bars represent 10 μm. (F) EM analyses of EXOC4-depleted HUVECs show immature WPBs with unfurled VWF (arrows). Scale bars represent 500 nm (i and iii) and 100 nm (ii and iv).