Investigating the molecular mechanisms of glycosaminoglycan assembly - Title: Investigating the molecular mechanisms of glycosaminoglycan assembly Project Summary The long-term goal of this research program is to elucidate and understand the regulatory mechanisms involved in the biosynthesis of glycosaminoglycans (GAGs) in mammalian cells. GAGs are long, linear polysaccharides that are expressed on all animal cells and play a key role in many cellular processes, including cell signaling and development. These complex carbohydrates are typically attached to core proteins, known as proteoglycans, located on the cell surface and in the extracellular matrix, and the chains are composed of alternating glucosamine and uronic acid sugar residues that are heterogeneously N- and O-sulfated. The biosynthesis of GAGs is a non-templated process, driven by the concerted activity of a large family of enzymes localized to the Golgi and endoplasmic reticulum. The arrangement and orientation of the sulfated sugar residues specify the location of distinct ligand binding sites on the cell surface, and these modifications can vary temporally during development and spatially across tissues. The capacity of GAGs to bind ligands impacts the fundamental properties of cells, the ability to form tissues and organs, and normal physiology. Despite the key functions of these molecules, there is a significant gap in knowledge regarding the regulatory mechanisms that give rise to their variable composition and binding properties. Through a multidisciplinary research program that leverages strengths in functional genomics, cell biology, and glycobiology, we aim to identify and characterize the mechanisms involved in controlling the inherent diversity of GAG structure and function in cells. Of particular interest, is the role of chromatin remodeling complexes in regulating GAG assembly during development and in disease states, as we recently identified members of the polycomb repressive complex (PRC) as novel epigenetic modifiers of GAG enzyme expression and assembly. We hypothesize that defined epigenetic and transcriptional programs tune the expression of biosynthetic enzymes in distinct cell types, which modulates their interaction with a plethora of growth factors and other binding partners in the extracellular matrix. We also aim to investigate how the core biosynthetic machinery is regulated in situ in the endoplasmic reticulum and Golgi. We plan to explore the physical association of the biosynthetic enzymes and core proteoglycans in the ER and Golgi and identify unknown chaperone and/or scaffolding proteins that may tune glycosylation in the secretory pathway. To carry out this work, we will leverage our historic strengths in the analysis of GAG structure, function, and regulation to understand: (1) how transcription factors and chromatin remodeling complexes control the expression of GAG biosynthetic enzymes, which impacts GAG structure and function, (2) how protein-protein interactions in the ER and Golgi orchestrate proteoglycan assembly in distinct cell types, and (3) how proteoglycan core proteins act as scaffolds for GAG assembly. Overall, we expect this endeavor to significantly advance our knowledge regarding the regulatory mechanisms controlling glycosylation and offer new strategies and targets to manipulate GAG biogenesis in human disease.
