Abstract 4587The nuclear proto-oncogene c-myb plays crucial roles in the growth, survival and differentiation of hematopoietic cells. We previously reported that the c-Myb knock down (KD) mice exhibited anemia and thrombocytosis. To better understand the transcriptional regulatory program that accompanies the decrease of c-Myb, we performed a transcriptome analysis of megakaryocyte-erythrocyte lineage-restricted progenitors (MEPs). We found that CD9 expression was dramatically increased in the MEPs from c-Myb KD mice. CD9 belongs to a tetraspanin family and regulates cell motility and adhesion. Several reports described that CD9 is involved in platelet function by forming a complex with integrin family members including platelet fibrinogen receptor integrin aIIb-b III. To elucidate the functional contribution of CD9 to the thrombogenesis within the regulatory hierarchy mediated by c-Myb, we first examine the relation between c-Myb and CD9. We found the recruitment of c-Myb to the promoter region of CD9 by chromatin immunoprecipitation assay. A reporter assay showed that c-Myb represses CD9 promoter activity. These results indicate that CD9 is directly repressed by c-Myb. Since an agonistic antibody against CD9 stimulated megakaryocytic colony formation, we investigated the role of CD9 during megakaryopoiesis in vivo by using CD9-null mice. Numbers of megakaryocytes and platelets, CFU-Meg, and ploidy were not different between wild-type and CD9-null mice. However, proplatelet formation (PPF) was significantly impaired in CD9-null megakaryocytes, and the size of proplatelets was smaller than those generated by wild-type megakaryocytes. Consistent with this result, the recovery phase of platelet counts were delayed in the CD9-null mice after the bone marrow suppression with 5-fluorouracil, indicating the impaired platelet production. In CD9-null mice, the proliferation of megakaryocytes was promoted and circulating megakaryocytes in the peripheral blood were increased, which might compensate the PPF impairment of CD9-null megakaryocytes. Our study suggests that c-Myb suppresses CD9 expression under the steady state, while, in the stress megakaryopoiesis, CD9 is derepressed and acts to enhance platelet production. Elucidation of c-Myb-CD9 regulatory function seems to be important to understand the stress megakaryopoiesis. Disclosures:Ninomiya:Alexion: Research Funding.