British Journal of Haematology , 2001, 113, 871±877 The storage defects in grey platelet syndrome and ad-storage pool deficiency affect a-granule factor V and multimerin storage without altering their proteolytic processing Catherine P. M. Hayward, 1,2 Harvey J. Weiss, 3 Bruce Lages, 3 Marisa Finlay, 1 Anna-Catharina Hegstad, 1 Shilun Zheng, 1 Alison Cowie, 1 Jean-Marc Masse Â, 4 Paul Harrison 5 and Elisabeth M. Cramer 4 Departments of 1 Pathology and Molecular Medicine and 2 Medicine, McMaster University and the Hamilton Regional Laboratory Medicine Program, Hamilton, Ontario, Canada, the 3 Department of Medicine St. Luke's/Roosevelt Hospital Center, Columbia University College of Physicians and Surgeons, New York, USA, 4 INSERM U474, Ho Ãpital de Port Royal, Paris, France, and 5 Haemostasis Research, Department of Haematology, University College Hospital, London, UK Received 9 November 2000; accepted for publication 21 February 2001 Summary. Among proteins stored in a-granules, multimerin and factor V share unusual features: they bind to each other, are proteolysed to unique forms and are stored eccentrically in a-granules. These unique features of their processing led us to study these proteins in alpha delta storage pool deficiency (ad-SPD) and grey platelet syndrome (GPS, a-SPD), two conditions known to impair a-granule protein storage. Platelet factor V and multimerin were severely reduced in GPS, whereas they ranged from reduced to normal in ad-SPD. The platelet levels of factor V and multimerin in these disorders indicated multimerin defi- ciency was not predictive of platelet factor V deficiency, although it reduced the amount of multimerin associated with platelet factor V. In GPS only, the defect in storing proteins was associated with increased multimerin and multimerin-factor V complexes in plasma. Like normal platelets, GPS and ad-SPD platelets contained factor V mainly in granules. Platelet factor V and multimerin were proteolysed to normal platelet forms in GPS and ad-SPD platelets, indicating that these conditions preserve some aspects of normal a-granule protein processing. Although we found factor V can be stored in platelets deficient in multimerin, our data indicate that multimerin storage influences the point at which multimerin binds factor V. Keywords: inherited platelet disorders, a-granules, multi- merin, factor V, platelet glycoproteins. Normal platelets store a large number of proteins in a- granules that they release during normal haemostasis (Harrison & Cramer, 1993). The processing and storage of a-granule proteins is thought to be important as a-granule protein deficiencies are associated with bleeding (reviewed in Hayward, 1997a). Although many proteins are processed for storage in a-granules, multimerin and factor V share some unique features. First, they undergo extensive proteolytic processing to unique forms found only in platelet a-granules (Viskup et al, 1987; Hayward, 1997b, 1999; Hayward et al, 1999). Second, they are stored as a non-covalent protein complex that sorts to an eccentric location within platelet a- granules independent of von Willebrand factor (the only other protein known to sort to this region) (Cramer et al, 1985a; Hayward et al, 1993, 1995; Chen et al, 1998). At present, the factors responsible for the unique processing of a-granule multimerin and factor V are not well understood. Unlike factor V, multimerin does not circulate in detectable amounts in normal plasma (Hay- ward, 1997b). When and where multimerin forms com- plexes with platelet factor V is not yet known (Hayward et al, 1995, 1999). Multimerin is processed for storage in secretion granules in megakaryocytes, endothelial cells and in heterologous cells capable of regulated secretion (Hayward et al, 1993, 1998, 1999). Megakaryocytes and endothelial cells express prepromultimerin transcripts of an identical size, however, promultimerin (M r 196 kDa, reduced) undergoes unique post-translational processing in megakaryocytes (Hayward, 1997b, 1999; Hayward et al, 1999). This generates the massive, variably sized q 2001 Blackwell Science Ltd 871 Correspondence: Dr Catherine P. M. Hayward, Room 2N32, McMaster University Medical Centre, 1200 Main St. W, Hamilton, Ontario, Canada L8N 3Z5. E-mail: haywrdc@mcmaster.ca