A single amino acid substitution in human Δpro restores association with canine VWFpp, enabling VWF storage.

In AtT-20 cells, Δpro's containing single amino acid substitutions were coexpressed in trans with canine VWFpp. Transfected cells were examined by confocal microscopy. 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 substitution of threonine at amino acid 869 to alanine in human Δpro, H-Thr869Ala-Δpro, resulted in a granular staining pattern for both canine VWFpp (A) and the mutated Δpro (B). The 2 proteins were colocalized in granules (C). The converse, an alanine-to-threonine substitution at amino acid 869 in canine Δpro (C-Ala869Thr-Δpro), resulted in a loss of storage of mutated VWF (D-F). Substitution of threonine 869 to proline in human VWF (H-Thr869Pro-Δpro) resulted in a loss of VWF storage (H), with normal granular storage of human VWFpp (G). These results indicate that amino acid 869 in VWF is important for the association with the VWFpp that results in VWF granular storage. Bar, 10 μm.