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 ß4GalT1, ß1 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 ß4GalT1 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 a2,6-
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.