The life-long production of blood cells requires hematopoietic developmental programs guided by systemic and local signals to convey the dynamic need for these cells. How these signals are delivered within the intra-medullar niches remain poorly understood. Extensive published and to-be-published information by the PIs of this Program, collaborative and individually, clearly established the involvement of glycans in the guidance of hematopoietic progenitor cell fate, function, and especially in thrombopoiesis and platelet functionality. The overarching hypothesis is ?cell-intrinsic and extrinsic glycan-mediated mechanisms regulate maintenance, differentiation, and function of hematopoietic cells.? Project 1 will investigate the roles of the galactosyltransferase ?4GalT1, ?1 integrin, and glycosaminoglycans (GAGs)/heparan sulfate proteoglycans (HSPGs) in thrombopoiesis at steady-state and following myeloablative stress using novel combined shared ?omics? and standard approaches with Project 3. A previously unknown role of ?4GalT1 to regulate megakaryocyte (MK) expression of HSPGs will also be investigated. A functionally defined MK-biased hematopoietic stem cell will be investigated together with Project 2, especially with respect to the heavily ?2,6- sialylated cell surface despite the absence of St6gal1 expression necessary to generate this structure. Project 2 will investigate the role of extracellular glycosylation, especially that mediated by extrinsic ST6GAL1 using the combined ?omics? approach (Project 1), how extrinsic sialylation in the hematopoietic niche is regulated, identifying the cell surface targets of sialylation; and with Project 3, understanding how the newly discovered GAG cofactor modulates extrinsic ST6GAL1 activity. Clinical Myelodysplastic Syndromes (MDS) and Myeloproliferative Neoplasms (MPN) and preclinical mouse models will be used in a first-time assessment into the glycobiology of these marrow diseases of highly heterogenous presentations but with the commonality of dysplastic MKs and altered platelet numbers and function, and analysis of these clinical diseases is shared across all three Projects. Project 3 will investigate the structure?function relationships of GAGs with proteins within the marrow microenvironment, such as growth factors and their receptors, and glycosyltransferases. Their roles in promoting thrombopoiesis and cell fate decisions will be interrogated using a multi-dimensional approach to identifying distinct GAG sequences. Project 3 will discover synthetic GAG mimetics as modulators of hematopoiesis/thrombopoiesis for therapeutic use. Core A will oversee the administration of the program, Core B will provide generation and sequencing of cDNA libraries derived from bulk RNA samples and single cells, and Core C will perform comparative structural analysis of GAGs, proteomics and protein-GAG interactions, and quantitative proteomics of protein expression. The three projects are intimately intertwined and will use all Cores. This program will uncover novel information to increase platelet production and help understand clinical conditions characterized by MK abnormalities.
The ability to produce blood cells is necessary throughout life, requiring hematopoietic developmental programs guided by cell-intrinsic and -extrinsic mechanisms conveying the need for these cells. The Program seeks to yield a transformative understanding of the role of glycosylation and glycans (cell surface sugars) in regulated blood cell production, in health and in disease. This knowledge will be leveraged to find novel therapeutic targets and therapeutics to promote hematopoietic health.
