The specific aims of this proposal test the central hypothesis that expression profiles of cellular glycans can serve as unique and sensitive fingerprints, capable of defining specific cell types or reporting cell status during culture. The development of markers and reagents for purifying specific sub-populations of cells is a high priority for advancing the therapeutic and investigational uses of hESCs. Despite their predominant localization to the cell surface, glycoconjugates have been under targeted for such uses. This deficiency reflects the lack of broadly accessible techniques or expertise for characterizing these complex macromolecules. We have developed a sophisticated suite of tools for quantitatively analyzing, with unprecedented depth, the glycans, glycoproteins, and glycolipids isolated from small amounts of material. We have successfully applied these tools to assess glycan expression changes associated with the differentiation of mouse ESCs and now propose to apply our developed glycomic and glycoproteomic techniques to questions of fundamental importance for human ESC biology. Our tools will allow us to screen for changes in glycosylation that occur globally or on specific molecules as a result of aneuploidy, cell culture conditions, passage number, cell pedigree, and differentiation.
In Specific Aim 1, we will define the glycomic fingerprints of hESC lines, and hESCs differentiated toward definitive endoderm, mesoderm, and neural precursor populations. The impact of aneuploidy and cell culture conditions will also be assessed.
In Specific Aim 2, we will map identified glycan markers, which define hESCs or differentiated cell fates, to specific sites on proteins and lipid cores.
In Specific Aim 3, we will generate tools for detecting and enriching sub-populations of hESCs and differentiated cells based on the expression of specific glycomic and/or glycoproteomic markers. Completion of these aims will not only provide novel approaches for characterizing, defining, and enriching specific cell types but will also provide a basis for exploring the role of glycosylation in hESC self renewal and differentiation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Program Projects (P01)
Project #
5P01GM085354-05
Application #
8382722
Study Section
Special Emphasis Panel (ZGM1-GDB-8)
Project Start
2012-08-01
Project End
2014-07-31
Budget Start
2012-08-01
Budget End
2013-07-31
Support Year
5
Fiscal Year
2012
Total Cost
$314,121
Indirect Cost
$101,158
Name
University of Georgia
Department
Type
DUNS #
004315578
City
Athens
State
GA
Country
United States
Zip Code
30602
Boward, Ben; Wu, Tianming; Dalton, Stephen (2016) Concise Review: Control of Cell Fate Through Cell Cycle and Pluripotency Networks. Stem Cells 34:1427-36
Foti, Rossana; Gnan, Stefano; Cornacchia, Daniela et al. (2016) Nuclear Architecture Organized by Rif1 Underpins the Replication-Timing Program. Mol Cell 61:260-73
Li, Ben; Sun, Zhaonan; He, Qing et al. (2016) Bayesian inference with historical data-based informative priors improves detection of differentially expressed genes. Bioinformatics 32:682-9
Rivera-Mulia, Juan Carlos; Gilbert, David M (2016) Replication timing and transcriptional control: beyond cause and effect-part III. Curr Opin Cell Biol 40:168-78
Avery, John; Dalton, Stephen (2016) Methods for Derivation of Multipotent Neural Crest Cells Derived from Human Pluripotent Stem Cells. Methods Mol Biol 1341:197-208
Rivera-Mulia, Juan Carlos; Gilbert, David M (2016) Replicating Large Genomes: Divide and Conquer. Mol Cell 62:756-65
Wilson, Korey A; Elefanty, Andrew G; Stanley, Edouard G et al. (2016) Spatio-temporal re-organization of replication foci accompanies replication domain consolidation during human pluripotent stem cell lineage specification. Cell Cycle 15:2464-75
Singh, Amar M; Trost, Robert; Boward, Benjamin et al. (2016) Utilizing FUCCI reporters to understand pluripotent stem cell biology. Methods 101:4-10
Berger, Ryan P; Sun, Yu Hua; Kulik, Michael et al. (2016) ST8SIA4-Dependent Polysialylation is Part of a Developmental Program Required for Germ Layer Formation from Human Pluripotent Stem Cells. Stem Cells 34:1742-52
Soufi, Abdenour; Dalton, Stephen (2016) Cycling through developmental decisions: how cell cycle dynamics control pluripotency, differentiation and reprogramming. Development 143:4301-4311

Showing the most recent 10 out of 97 publications