One of the signaling pathways that play pivotal roles in stem cell and hematopoietic homeostasis is the Notch signaling cascade. Notch plays a critical role in many developmental and pathophysiological processes. Mutation or abnormal activation of Notch pathway is linked to a variety of human development disorders and cancers. Notch receptor modification by a sugar molecule, fucose, critically regulates Notch receptor interaction with their counterparts, Notch ligands, and is important for Notch signaling activation. The addition of fucose to Notch receptors are mediated by protein O-fucosyltransferase 1 (Pofut1). Using two mouse models, in which mice are genetically modified to be conditionally deficient either in the expression of all fucose-carrying glycoproteins or glycolipids (FX-/-), or only O-linked fucosyl-glycans present on EGF repeats of a few proteins including Notch (conditional Pofut1-/-), we recently identified a role of O-fucose-modified Notch signaling that is essential for hematopoietic homeostasis as well as a requirement of fucose modification of hematopoietic stem cell (HSC) for their proper location in the bone marrow microenvironment, also called niche. These findings uncovered a unique role of fucose-modified Notch in hematopoiesis and HSC biology. In this study, in order to test the hypothesis that fucose-modified Notch is critical for Notch-ligand interaction- dependent control of HSC homeostasis, we will first define the impact of lack of Notch fucose moieties that are required for ligand binding on HSC homeostasis and blood lineage differentiation by using genetically modified mouse models and mouse embryonic stem cells. We will also assess if Pofut1-deficient HSCs, or HSCs that lack fucose known essential for Notch ligand binding, are displaced in the proper marrow niches by using the cutting edge imaging technology (2-photon intravital microscopy). Then, we will determine the mechanism(s) by which lack of Notch O-fucose causes aberrant HSC homeostasis. We will assess if a disruptive interaction between Notch and its ligand due to the lack of fucose in the ligand binding domain is the main reason that causes aberrant stem cell niche location and abnormal HSC function. Further, we will assess the interaction between Notch pathway with other major adhesion molecules and cytokines that are important for stem cell bone marrow homing and marrow niche lodging. Finally, we will determine the significance of the enhanced fucose modification of Notch in HSC binding with ligand and in HSC marrow homing and lodgment. At the end of these studies, we hope the knowledge we have gained will improve our understanding of the role of Notch glycosylation in stem cell biology, and will form the basis that leads to future studies and development of novel therapeutic maneuvers in stem cell therapy and cancer medicine.

Public Health Relevance

HSC functions are intimately regulated by a coordinated interaction of a variety of external cues and intrinsic factors. This study will facilitate our understanding of the mechanisms of Notch signaling in HSC homeostasis and in leukemic stem cell biology, and the importance of Notch fucosylation in HSC niche competency and HSC functional control. These findings will allow us to design novel approaches in stem cell transplantation and stem cell therapy in the future.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL103827-02
Application #
8399085
Study Section
Molecular and Cellular Hematology (MCH)
Program Officer
Thomas, John
Project Start
2011-12-15
Project End
2016-11-30
Budget Start
2012-12-01
Budget End
2013-11-30
Support Year
2
Fiscal Year
2013
Total Cost
$373,660
Indirect Cost
$135,660
Name
Case Western Reserve University
Department
Pathology
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Murata, Akihiko; Yoshino, Miya; Hikosaka, Mari et al. (2014) An evolutionary-conserved function of mammalian notch family members as cell adhesion molecules. PLoS One 9:e108535