The processes of megakaryocyte differentiation, proplatelet formation, and the daily release of 1011 platelets into the bloodstream are tightly regulated. Genetic disturbances can lead to a cascade of downstream molecular alterations that markedly affect the function of megakaryocytes and platelets. Therefore, identifying new genes and their function in megakaryocytes and platelets is critical for understanding how these unique cells contribute to health and disease. Over the last decade advances in genomics, specifically next generation sequencing, have allowed for the discovery of several mutations and genetic variants that cause disease or influence associated hematological traits. By performing platelet RNA-Seq we were among the first to identify NBEAL2 as the causative gene for gray platelet syndrome (GPS) and showed that NBEAL2 regulates megakaryocyte development and platelet function.1-3 Mice carrying targeted Nbeal2 null alleles not only replicated the thrombocytopenia and lack of alpha granules observed in humans, but also provided new information about the role of platelets in thromboinflammation, wound healing, myelofibrosis and metastasis dissemination.4-7 More recently, we and others found that germline mutations in ETV6 lead to thrombocytopenia, red cell macrocytosis, and predisposition to lymphoblastic leukemia.8,9ETV6 encodes an ETS family transcriptional repressor, which exerts its activity by binding a consensus sequence in the promoter regions of DNA. Mice with conditional Etv6 knockout in megakaryocytic-erythroid cells are thrombocytopenic indicating the involvement of Etv6 in thrombopoiesis.10 Several of the families recently described have a missense mutation in the central domain of ETV6 (p.P214L). This mutation results in aberrant cellular localization of ETV6, decreased transcriptional repression, and impaired megakaryocyte maturation. The bone marrow of individuals affected by this mutation show hyperplasia of immature megakaryocytes suggesting a differentiation arrest. Deep sequencing of the platelet transcriptome also revealed significant differences in mRNA expression levels between patients with the ETV6 p.P214L mutation and non-affected family members, indicating that ETV6 is critically involved in defining the molecular phenotype and function of platelets. Consistent with this notion, individuals with the ETV6 p.P214L mutation experience bleeding that is disproportionate to their mild thrombocytopenia. We have also used CRISPR/Cas9 technology to generate a mouse colony where the human p.P214L ETV6 mutation was inserted into the conserved site of Etv6. Mice with this mutation (Etv6H.P214L) have reduced platelet counts. In summary, advances in human genetics that led to the discovery of novel congenital platelet disorders coupled with relevant animal models will likely contribute to our understanding of megakaryopoiesis and platelet function.
References
1. Kahr WH, Hinckley J, Li L, et al. Mutations in NBEAL2, encoding a BEACH protein, cause gray platelet syndrome. Nature genetics. 2011;43(8):738-740.
2. Gunay-Aygun M, Falik-Zaccai TC, Vilboux T, et al. NBEAL2 is mutated in gray platelet syndrome and is required for biogenesis of platelet alpha-granules. Nature genetics. 2011;43(8):732-734.
3. Albers CA, Cvejic A, Favier R, et al. Exome sequencing identifies NBEAL2 as the causative gene for gray platelet syndrome. Nature genetics. 2011;43(8):735-737.
4. Deppermann C, Cherpokova D, Nurden P, et al. Gray platelet syndrome and defective thrombo-inflammation in Nbeal2-deficient mice. The Journal of clinical investigation. 2013.
5. Kahr WH, Lo RW, Li L, et al. Abnormal megakaryocyte development and platelet function in Nbeal2(-/-) mice. Blood. 2013;122(19):3349-3358.
6. Guerrero JA, Bennett C, van der Weyden L, et al. Gray platelet syndrome: proinflammatory megakaryocytes and alpha-granule loss cause myelofibrosis and confer metastasis resistance in mice. Blood.2014;124(24):3624-3635.
7. Tomberg K, Khoriaty R, Westrick RJ, et al. Spontaneous 8bp Deletion in Nbeal2 Recapitulates the Gray Platelet Syndrome in Mice. PLoS One. 2016;11(3):e0150852.
8. Noetzli L, Lo RW, Lee-Sherick AB, et al. Germline mutations in ETV6 are associated with thrombocytopenia, red cell macrocytosis and predisposition to lymphoblastic leukemia. Nature Genetics. 2015;47(5):535-538.
9. Zhang MY, Churpek JE, Keel SB, et al. Germline ETV6 mutations in familial thrombocytopenia and hematologic malignancy. Nature genetics. 2015;47(2):180-185.
10. Wang LC, Swat W, Fujiwara Y, et al. The TEL/ETV6 gene is required specifically for hematopoiesis in the bone marrow. Genes & development. 1998;12(15):2392-2402.
Disclosures
Di Paola: CSL BEhring: Consultancy; Biogen: Consultancy.
Megakaryocytic emperipolesis and platelet function abnormalities in five patients with gray platelet syndrome