PROJECT SUMMARY/ ABSTRACT Congenital disorders of glycosylation are multi-system inherited diseases, which affect the amount of glycosylation added to proteins. For patients with mannose phosphate isomerase (MPI) deficiency, decreased mannose production leads to globally decreased levels of N-linked glycans resulting in several issues with congenital diarrhea and protein losing enteropathy being the most significant symptoms. Our long-term objective is to determine how defective glycosylation manifests as disease. We aim to achieve this objective using a novel mouse strain named benadryl, which has an Mpi mutation and models most of the features of human disease. We found conditional knockout of Mpi in the intestines re-created cardinal features of the benadryl strain with severe defects in the mucus producing goblet cells, and features of inflammatory bowel disease. The goal of this project is to determine how Mpi protects the intestines and how to improve therapeutic interventions for patients deficient in Mpi. The central hypothesis is that Mpi is a rate limiting step for N- glycosylation of mucins like Muc2, which are essential for proper mucin maturation and barrier function. In this proposal we will investigate (Aim 1) how N-glycosylation of Muc2 is required for goblet cell survival, intestinal mucus production, and protection from bacteria; (Aim 2) how N-glycosylation sites are essential for proper Muc2 folding and dimerization; and (Aim 3) the mechanism for how mannose therapy protects gastrointestinal barriers. Upon finishing these studies, we will have defined the role of N-glycosylation post-translation modification of Muc2 in maintaining a proper intestinal mucus barrier to prevent bacterial invasion and inflammation. Further, we will leverage N-glycosylation insights to test feasibility and mechanism of mannose as a therapy in pre-clinical models of gastrointestinal disease.