Time-Resolved Interactome Profiling Deconvolutes Secretory Protein Quality Control Dynamics

Abstract

SUMMARYMany cellular processes are governed by protein-protein interactions that require tight spatial and temporal regulation. Accordingly, it is necessary to understand the dynamics of protein interactions to fully comprehend and elucidate cellular processes and pathological disease states. To map de novo protein-protein interactions with time-resolution at an organelle-wide scale, we developed a quantitative mass-spectrometry method, time-resolved interactome profiling (TRIP). We apply TRIP to elucidate aberrant protein interaction dynamics that lead to the protein misfolding disease congenital hypothyroidism. We deconvolute altered temporal interactions of the thyroid hormone precursor thyroglobulin with pathways associated with hypothyroidism pathophysiology such as Hsp70/90 assisted folding, disulfide/redox processing, and N-glycosylation. Functional siRNA screening identified VCP and TEX264 as key protein degradation components whose inhibition selectively rescues mutant prohormone secretion. Ultimately, our results provide insight into the temporal coordination of protein homeostasis, and our TRIP method can have broad application in investigating protein folding diseases and cellular processes.HIGHLIGHTSMethod to map de novo protein-protein interactions with temporal resolutionCharacterization of the time-resolved interactome of a protein folding disease statesiRNA screening identifies VCP and TEX264 as major regulators of thyroglobulin processingVCP pharmacological inhibition rescues mutant thyroglobulin secretion