| dc.description.abstract | The endoplasmic reticulum (ER), responsible for processing approximately one-third of the human proteome including most secreted and membrane proteins, plays a pivotal role in protein homeostasis (proteostasis). Dysregulation of ER proteostasis is implicated in a number of disease states. As such, continued efforts are directed at elucidating mechanisms of ER protein quality control mediated by transient and dynamic protein-protein interactions with molecular chaperones, co-chaperones, protein folding and trafficking factors that take place in and around the ER. Thyroglobulin (Tg) is a secreted iodoglycoprotein serving as the precursor for T3 and T4 hormones. Many characterized Tg gene mutations produce secretion-defective variants resulting in congenital hypothyroidism (CH). While dependencies on individual proteostasis network components were known for Tg, the integration of proteostasis pathways mediating Tg protein quality control and the molecular basis of mutant Tg misprocessing remained poorly understood. Interaction changes between WT and several CH-variants are associated with common imbalances in proteostasis engagement including increased chaperoning, oxidative folding, and engagement by targeting factors for ER-associated degradation (ERAD). Furthermore, mutation-specific changes in engagement with N-glycosylation components reveal unique dynamics for degradation. These interactions require tight spatial and temporal regulation. Accordingly, understanding these parameters within the dynamics of protein quality control is necessary to fully comprehend and elucidate cellular processes and pathological disease states associated with them. Time-resolved interactome profiling (TRIP) allows for mapping de novo protein-protein interactions with time-resolution at an organelle-wide level. TRIP allows for the deconvolution of altered temporal engagement of Tg across secretory proteoastasis pathways such as Hsp70/90 assisted folding, disulfide/redox processing, and N-glcosylation associated with CH pathophysiology. Functional siRNA screening identified key protein degradation components whose inhibition selectively rescues mutant Tg secretion. Ultimately, these data provide insight into the temporal coordination of secretory proteostasis and may have broad application in investigating other protein folding diseases and cellular processes. | |
