CLATHRIN MEDIATED ENDOCYTOSIS IS REGULATED BY THE NUTRIENT SENSOR O-GlcNAc TRANSFERASE

Candidate: Sadia Rahmani
Supervisor: Dr. Costin Antonescu and Dr. Warren Wakarchuk

ABSTRACT

Clathrin-mediated endocytosis (CME) is a multifunctional biological process involved in internalization of receptors, viruses, and nutrients from the cell surface. CME has important implications for different cell surface proteins such as Receptor Tyrosine Kinases that regulate growth, adhesion, and migration. In turn, CME is highly regulated to control the abundance of cell surface proteins under different physiological requirements of the cell. I examined how the modification of proteins with O-linked N-acetyl-glucosamine (O-GlcNAc) may control CME. I used perturbation of key enzymes responsible for protein O-GlcNAc modification, as well as specific mutants of the central endocytic regulator. I identified that initiation and the assembly of clathrin and other proteins is controlled by O-GlcNAc protein modification. This reveals a new dimension of regulation of CME and highlights the important reciprocal regulation of cellular metabolism and endocytosis.

The dynamic O-GlcNAc modification is added by O-GlcNAc transferase (OGT) and removed by O-GlcNAc hydrolase within nucleus and cytoplasm. OGT is an emergency nutrient sensor amongst others such as AMP-Activated Protein Kinase (AMPK) that may fine-tune cellular responses to stress stimuli. I define the metabolic conditions in which OGT and AMPK are reciprocally regulated thus revealing the cooperativity and signaling crosstalk between the nutrient sensors. 

Having established a key role for the dynamic O-GlcNAc modification, I also explored the synthetic modification of cell surface glycoproteins with a stable and elaborate polymer of the sugar sialic acid. Using a recombinant bacterial enzyme for synthesis of poly-α2,8-N-acetylneuraminic acid (polysialic acid; polySia) onto cardiomyocytes increased their propensity for persistent directional cellular migration. As such, my work characterizes the cellular effects of synthetic PolySia for cellular therapy.