Mapping and Exploring the Collagen-I Proteostasis Network
Author(s)
DiChiara, Andrew Stephen; Taylor, Rebecca J.; Wong, Madeline Y.; Doan, Ngoc Duc; Del Rosario, Amanda M; Shoulders, Matthew D.; ... Show more Show less
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Collagen-I is the most abundant protein in the human body, yet our understanding of how the endoplasmic reticulum regulates collagen-I proteostasis (folding, quality control, and secretion) remains immature. Of particular importance, interactomic studies to map the collagen-I proteostasis network have never been performed. Such studies would provide insight into mechanisms of collagen-I folding and misfolding in cells, an area that is particularly important owing to the prominence of the collagen misfolding-related diseases. Here, we overcome key roadblocks to progress in this area by generating stable fibrosarcoma cells that inducibly express properly folded and modified collagen-I strands tagged with distinctive antibody epitopes. Selective immunoprecipitation of collagen-I from these cells integrated with quantitative mass spectrometry-based proteomics permits the first mapping of the collagen-I proteostasis network. Biochemical validation of the resulting map leads to the assignment of numerous new players in collagen-I proteostasis, and the unanticipated discovery of apparent aspartyl-hydroxylation as a new post-translational modification in the N-propeptide of collagen-I. Furthermore, quantitative analyses reveal that Erp29, an abundant endoplasmic reticulum proteostasis machinery component with few known functions, plays a key role in collagen-I retention under ascorbate-deficient conditions. In summary, the work here provides fresh insights into the molecular mechanisms of collagen-I proteostasis, y ielding a detailed roadmap for future investigations. Straightforward adaptations of the cellular platform developed will also enable hypothesis-driven, comparative research on the likely distinctive proteostasis mechanisms engaged by normal and disease-causing, misfolding collagen-I variants, potentially motivating new therapeutic strategies for currently incurable collagenopathies.
Date issued
2016-02Department
Massachusetts Institute of Technology. Department of Chemistry; Koch Institute for Integrative Cancer Research at MITJournal
ACS Chemical Biology
Publisher
American Chemical Society (ACS)
Citation
DiChiara, Andrew S. et al. “Mapping and Exploring the Collagen-I Proteostasis Network.” ACS Chemical Biology 11, 5 (March 2016): 1408–1421 © 2016 American Chemical Society
Version: Author's final manuscript
ISSN
1554-8929
1554-8937