Glycans decorate eukaryotic cell surface and secreted proteins, where they are poised to mediate a variety of molecular recognition events involved in attachment viruses and bacterial infection, immune cell activation, cancer metastasis and organ development. Many biological functions of cell-surface glycans can only be studied in the context of living cells. In addition to a glycan's primary structure, the architecture of its protein scaffold, as well as its density and distribution on the cell surface can all contribute to biological activity. Unfortunately, the heterogeneous nature of cell surfaces, particularly with respect to glycoconjugate structures, has frustrated molecular-level studies of glycan function. The controlled presentation of structurally defined glycoconjugates on live cells is therefore an important capability for accelerating research in glycobiology. The broad objective of this project is to develop chemical approaches for engineering cell surface glycan structures. During the previous granting period, we developed a method for controlling the activity of glycan biosynthetic enzymes in the Golgi compartment using the chemical inducer of dimerization strategy. We engineered Golgi sulfotransferases and glycosyltransferases for control by chemical dimerizers and demonstrated that cell surface expression of selectin ligands and blood group antigens can be modulated accordingly. We also developed a chemical dimerizer for applications in animal models. The next granting period we will focus three specific Aims. First, we will develop a method for site- specific chemical modification of cell surface proteins with synthetic, structurally-defined glycans. The method employs the 6-residue consensus sequence for formylglycine generating enzyme (FGE), which drives the formation of formylglycine (FGly) from a genetically encoded cysteine residue. The aldehyde functionality of FGly provides a uniquely reactive site to which synthetic aminooxy glycans can be ligated via oxime formation. We will use the """"""""aldehyde tag"""""""" to modify recombinant cell surface proteins with tailored glycans. Applications to the construction of artificial cell surface mucins and to studies of the P-selectin ligand PSGL-1 are proposed.
The second Aim i nvolves application of the aldehyde tag to biophysical studies of the natural cell-surface mucin MUC1. The third and final Aim focuses on a second technology for engineering cell surface glycans. We plan to develop synthetic polymers that mimic mucin glycoproteins and to study their behavior when inserted into live cell membranes. We will investigate their cell surface dynamics and membrane orientations. The synthetic mucin mimics will then be applied to studies of siglec recognition and signaling.
Complex sugars decorate the surfaces of cells, where they play a role in cell-cell interactions involved in normal biological processes and also in human disease. The goal of this research is to develop chemical tools for studying the functions of complex sugars on the cell surface. These tools will improve our understanding of how sugars contribute to diseases such as cancer and inflammation.
|Barnes, J Matthew; Przybyla, Laralynne; Weaver, Valerie M (2017) Tissue mechanics regulate brain development, homeostasis and disease. J Cell Sci 130:71-82|
|Freeman, Spencer A; Goyette, Jesse; Furuya, Wendy et al. (2016) Integrins Form an Expanding Diffusional Barrier that Coordinates Phagocytosis. Cell 164:128-140|
|Bhat, Ramray; Belardi, Brian; Mori, Hidetoshi et al. (2016) Nuclear repartitioning of galectin-1 by an extracellular glycan switch regulates mammary morphogenesis. Proc Natl Acad Sci U S A 113:E4820-7|
|Hudak, Jason E; Belardi, Brian; Appel, Mason J et al. (2016) Piperidine-based glycodendrons as protein N-glycan prosthetics. Bioorg Med Chem 24:4791-4800|
|Kai, FuiBoon; Laklai, Hanane; Weaver, Valerie M (2016) Force Matters: Biomechanical Regulation of Cell Invasion and Migration in Disease. Trends Cell Biol 26:486-497|
|Xiao, Han; Woods, Elliot C; Vukojicic, Petar et al. (2016) Precision glycocalyx editing as a strategy for cancer immunotherapy. Proc Natl Acad Sci U S A 113:10304-9|
|Agarwal, Paresh; Beahm, Brendan J; Shieh, Peyton et al. (2015) Systemic Fluorescence Imaging of Zebrafish Glycans with Bioorthogonal Chemistry. Angew Chem Int Ed Engl 54:11504-10|
|Agarwal, Paresh; Bertozzi, Carolyn R (2015) Site-specific antibody-drug conjugates: the nexus of bioorthogonal chemistry, protein engineering, and drug development. Bioconjug Chem 26:176-92|
|Woods, Elliot C; Yee, Nathan A; Shen, Jeff et al. (2015) Glycocalyx Engineering with a Recycling Glycopolymer that Increases Cell Survival In Vivo. Angew Chem Int Ed Engl 54:15782-8|
|Kramer, Jessica R; Onoa, Bibiana; Bustamante, Carlos et al. (2015) Chemically tunable mucin chimeras assembled on living cells. Proc Natl Acad Sci U S A 112:12574-9|
Showing the most recent 10 out of 59 publications