Intracellular localization of chimeric Δpro VWF coexpressed in trans with canine VWFpp.

AtT-20 cells were transiently cotransfected with canine VWFpp and human/canine chimeric Δpro to determine what portion of VWF must contain canine sequence to restore association and granular sorting of VWF. Panels A, D, and G show cells stained for VWFpp (red). Panels B, E, and H show cells stained for VWF (green). The merges of VWFpp and VWF staining are shown in panels C, F, and I. The chimeric Δpro, H/C-866-Δpro, was coexpressed with canine VWFpp and demonstrated a granular staining pattern for both VWFpp (A) and the chimeric Δpro (B). The 2 proteins were colocalized in granules (C). The converse chimeric Δpro, C/H-866-Δpro, was also coexpressed with canine VWFpp; it demonstrated a granular staining pattern for VWFpp (D) but only a diffuse staining pattern for VWF (E). Canine VWFpp was coexpressed with a third chimeric Δpro, C/H-907-Δpro. Both the canine VWFpp (G) and chimeric Δpro (H) were stored in granules, where they colocalized (I). When the first 143 amino acids of mature VWF contain canine sequence, association with canine VWFpp is restored and results in granular sorting of VWF. Bar, 10 μm.