News and Publications

Transgenic Models of Platelet Structure and Function

J. Ware, J. Dent, C. Martinez, K. Kato, S. Russell, T. Kanaji

A PROTOTYPIC PLATELET SEPTIN AND ITS PARTICIPATION IN SECRETION

Septins are a family of proteins originally identified in yeasts as essential for budding. For years, the presence of septins in higher eukaryotic cells was dismissed because budding, or asymmetric cell division, is a process unique to yeasts. However, recent studies indicated that septins do exist in higher eukaryotes. In those cases examined, eukaryotic septins were associated with events in which dynamic membrane movement or cytoplasmic partitioning, as in cytokinesis or vesicle trafficking, occurs.

In characterizing transcripts from the human chromosome 22q11.2 locus, we identified and characterized a human septin gene, which we termed CDCREL-1 (cell division control related-1). The highest levels of CDCrel-1, the gene product, are found in neurons and circulating blood platelets. The results of cloning studies of CDCREL-1 suggested that it might be involved in the release of neurotransmitters.

Some molecular similarities exist between platelet secretion and the mechanisms that control neurotransmitter release. Although the mechanisms that control release of neurotransmitters have been intensively studied, the molecular basis of platelet secretion remains poorly characterized. We found that platelet CDCrel-1 is a protein kinase substrate in the presence of known platelet agonists. Immunopurification of CDCrel-1 revealed that it is part of a macromolecular complex containing a protein involved in platelet secretion, syntaxin 4. We localized CDCrel-1 to areas surrounding platelet storage granules.

To determine the relevance of CDCrel-1 to normal platelet function, we generated mice that lacked CDCREL-1 and characterized platelets from these animals. Compared with platelets from wild-type littermates, platelets from the mutant mice aggregated and released stored serotonin in the presence of subthreshold levels of collagen. Our results provide new insights into the mechanisms that regulate platelet secretion and indicate that platelet septins contribute to membrane trafficking within megakaryocytes and platelets.

MURINE MODELS OF BERNARD-SOULIER SYNDROME

Disorders characterized by giant platelets are a heterogeneous group of hematopoietic defects. Many of these disorders are hereditary and are associated with reduced platelet counts, findings that lead to their classification as hereditary macrothrombocytopenias. The molecular pathologic changes leading to macrothrombocytopenia are obscure but most likely are linked to abnormal megakaryocyte maturation and platelet release.

The Bernard-Soulier syndrome is one of the best known syndromes among those characterized by macrothrombocytopenia. The syndrome is due to mutations within any of the 3 subunits that make up the glycoprotein Ib-IX complex (GPIb-IX), a membrane receptor. Most commonly, mutations in GPIb-IX prevent translocation of the receptor to the megakaryocyte surface, resulting in the Bernard-Soulier syndrome. The bleeding associated with the syndrome is more severe than would be expected on the basis of the macrothrombocytopenia and reflects the critical role for GPIb-IX in platelet adhesion during hemostasis. Indeed, the role of GPIb-IX in hemostasis is well established, but the molecular basis for the macrothrombocytopenia is unknown.

We recently described a murine model of human Bernard-Soulier syndrome in which the mice have all of the important features associated with the human syndrome. Our studies on bone marrow from mice that lack the gene for GPIb suggest that mature megakaryocytes have a reduced quantity of membrane and do not form platelet fields in the mature megakaryocytes, the precursor cells of platelets. The aberrant membrane development leads to abnormal fragmentation in the vascular sinus and abnormal proplatelets. The use of mouse models to examine platelet production is providing novel insights into the structural elements of GPIb-IX that control normal platelet genesis and function.

PUBLICATIONS

Dent, J., Kato, K., Peng, X.R., Martinez, C., Cattaneo, M., Poujol, C., Nurden, P., Nurden, A., Trimble, W.S., Ware, J. A prototypic platelet septin and its participation in secretion. Proc. Natl. Acad. Sci. U. S. A. 99:3064, 2002.

Kasirer-Friede, A., Ware, J., Leng, L., Marchese, P., Ruggeri, Z.M., Shattil, S.J. Lateral clustering of platelet GP Ib-IX complexes leads to up-regulation of the adhesive function of integrin a_IIbb₃. J. Biol. Chem. 277:11949, 2002.

Peng, X.R., Jia, Z., Zhang, Y., Ware, J., Trimble, W.S. The septin CDCrel-1 is dispensable for normal development and neurotransmitter release. Mol. Cell. Biol. 22:378, 2002.

Poujol, C., Ware, J., Nieswandt, B., Nurden, A.T., Nurden, P. Absence of GPIb is responsible for aberrant membrane development during megakaryocyte maturation: ultrastructural study using a transgenic model. Exp. Hematol. 30:352, 2002.

Ware, J., Ruggeri, Z.M. Platelet receptors: von Willebrand factor. In: Platelets in Thrombotic and Non-Thrombotic Disorders. Gresele, P., et al. (Eds.). Cambridge University Press, New York, 2002, p. 111.