SEQUENCING THE MHC CLASS I PEPTIDES FROM TRYPANOSOMA CRUZI

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DESCRIPTION (provided by applicant): Glycoproteins and glycolipids have many functions in human physiology and disease and the expression of the glycans on these cell surface molecules are known to change during early embryonic development. Embryonic stem cells, derived from early embryos, can be grown in vitro and induced to differentiate into many cell types. Controlling this differentiation and producing homogeneous populations of differentiated stem cells are major challenges for the biomedical community. Development of cell markers and technology to enrich cell populations could aid significantly stem cell research. We propose to establish an Integrated Resource Center for Biomedical Glycomics that will focus on developing tools and technology to analyze in detail the glycoprotein and glycolipid expression of mouse embryonic stem cells and cells into which they differentiate. Project 1, Embryonic Stem Cell Program, will develop the tools to culture large numbers of stem cells and investigate in detail the glycans expressed on specific cell surface glycoproteins. Project 2 will develop technology to fractionate glycopeptides produced from stem cells and determine the parent proteins that express these glycans and at which sites these glycans are expressed. Project 3 will develop kinetic real-time PCR to study the expression of glycosyltransferase transcripts. Project 4 will develop Bioinformatic tools to input structural and expression data into relational databases for processing and analysis. The technology developed in this Resource will allow an understanding of how glycosylation is controlled during differentiation and will allow the development of tools to promote the use of stem cells to treat human disease. In addition, the technology will be applicable to the study of other cell types, including cancer cells that are progressing to a more invasive phenotype. This glycomics resource will result in technology development that will allow others in the scientific community to participate in glycomics research via dissemination of the new methods developed and via analytical service provided by the resource to other scientists requesting help in glycomic analyses. Training courses in the developed technologies will also be provided to the scientific community. Collaborative projects already constitute a significant amount of the proposed research and will become an increasingly important component of the developing glycomics resource.