Despite tremendous advances in our understanding of platelet physiology, the underlying molecular mechanisms remain unknown in the vast majority of patients with inherited defects in platelet function. Mechanisms, such as deficiencies in membrane glycoproteins and the storage pool deficiency, are commonly considered but occur in a small proportion of patients. Most of these patients are characterized by a bleeding disorder, and impaired aggregation and dense granule secretion on platelet activation, and are currently lumped in a loosely defined group called "platelet secretion/activation defects." The molecular mechanisms in these patients are unknown. Several studies suggest that these patients may have abnormalities in the signal transduction mechanisms. We have documented specific defects in platelet phospholipase C-beta2, Gaq and protein kinase C-theta. Newer approaches are required to define the molecular defects in such patients who constitute an untapped reservoir of new information. We have now applied genome-wide platelet expression profiling using the Affymetrix microarrays to demonstrate a unique decrease in expression of myosin light chain 9 (MYL9, 70 fold decrease) and other genes in a patient with an inherited thrombocytopenia, impaired platelet responses (including myosin and pleckstrin phosphorylation and GPIIb-llla activation), and a heterozygous mutation in transcription factor CBFA2 (Core-binding factor A2), thereby providing proof of concept that expression profiling can indeed be applied to obtain new insights in patients with platelet dysfunction. The microarray technology has hitherto not been applied to such patients. In this project, we propose an integrated approach encompassing, a) detailed biochemical and functional studies on platelet mechanisms, b) platelet expression profiling and c) subsequent validation of the aberrant mechanisms using functional, mutational, biochemical and immunological approaches, to delineate the molecular mechanisms in 15 patients with inherited platelet dysfunction. We have extensive experience in all of the involved approaches. This project is a collaborative effort between two laboratories with strong complementary expertise. It represents application of the recent advances in genomics and the Human Genome Project to a group of patients who are frequently encountered but extremely poorly characterized. Our studies will provide a wealth of information on platelet mechanisms and on potential new targets for antithrombotic therapy.