INTRACELLULAR TRAFFICKING OF FACTOR-VIII TO VON-WILLEBRAND-FACTOR STORAGE GRANULES

In plasma, von Willebrand factor (vWf) associates with Factor VIII (FV III); however, the site at which these proteins first interact has not been defined, Administration of 1-desamino-8-D-arginine vasopressin ( DDAVP) causes a rapid, concomitant elevation in plasma levels of both vWf and FVIII, suggesting the existence of a DDAVP-releasable storage pool for both proteins, To determine whether vWf and FVIII can associa te intracellularly and colocalize to storage vesicles, we transfected AtT-20 cells with vWf and FVIII expression plasmids, FVIII alone was n ot detectable within storage granules; however, transfection of vWf cD NA into the same cell caused FVIII to alter its intracellular traffick ing and to undergo granular storage, colocalizing to the vWf-containin g granules, In contrast, colocalization of FVIII was not observed when these cells were transfected with plasmids encoding defective FVIII-b inding vWf mutants, Transfection of bovine endothelial cells with FVII I further demonstrated vesicular storage of FVIII with vWf in Weibel-P alade bodies. Since gene therapy of hemophilia A may ultimately target endothelium or hematopoietic stem cells, the interaction between vWf and FVIII within a secretory cell is important, Thus, vWf can alter th e intracellular trafficking of FVIII from a constitutive to a regulate d secretory pathway, thereby producing an intracellular storage pool o f both proteins.