Mapping and Exploring the Collagen-I Proteostasis Network
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 ha...
| Main Authors: | , , , , , |
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| Format: | Article |
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American Chemical Society (ACS)
2018
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| Online Access: | http://hdl.handle.net/1721.1/113577 https://orcid.org/0000-0002-9438-0643 https://orcid.org/0000-0002-0508-5002 https://orcid.org/0000-0001-9672-2064 https://orcid.org/0000-0002-6511-3431 |
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| author | DiChiara, Andrew Stephen Taylor, Rebecca J. Wong, Madeline Y. Doan, Ngoc Duc Del Rosario, Amanda M Shoulders, Matthew D. |
| author2 | Massachusetts Institute of Technology. Department of Chemistry |
| author_facet | Massachusetts Institute of Technology. Department of Chemistry DiChiara, Andrew Stephen Taylor, Rebecca J. Wong, Madeline Y. Doan, Ngoc Duc Del Rosario, Amanda M Shoulders, Matthew D. |
| author_sort | DiChiara, Andrew Stephen |
| collection | MIT |
| description | 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. |
| first_indexed | 2024-09-23T14:52:08Z |
| format | Article |
| id | mit-1721.1/113577 |
| institution | Massachusetts Institute of Technology |
| last_indexed | 2024-09-23T14:52:08Z |
| publishDate | 2018 |
| publisher | American Chemical Society (ACS) |
| record_format | dspace |
| spelling | mit-1721.1/1135772022-09-29T11:05:59Z Mapping and Exploring the Collagen-I Proteostasis Network DiChiara, Andrew Stephen Taylor, Rebecca J. Wong, Madeline Y. Doan, Ngoc Duc Del Rosario, Amanda M Shoulders, Matthew D. Massachusetts Institute of Technology. Department of Chemistry Koch Institute for Integrative Cancer Research at MIT DiChiara, Andrew Stephen Taylor, Rebecca J. Wong, Madeline Y. Doan, Ngoc Duc Del Rosario, Amanda M Shoulders, Matthew D. 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. National Institute of Arthritis and Musculoskeletal and Skin Diseases (U.S.) (Grant 1R03AR067503) National Institute of Arthritis and Musculoskeletal and Skin Diseases (U.S.) (Grant 1F31AR067615) National Institute of Environmental Health Sciences (Grant P30-ES002109) 2018-02-12T16:15:03Z 2018-02-12T16:15:03Z 2016-02 2015-12 2018-02-06T18:35:29Z Article http://purl.org/eprint/type/JournalArticle 1554-8929 1554-8937 http://hdl.handle.net/1721.1/113577 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 https://orcid.org/0000-0002-9438-0643 https://orcid.org/0000-0002-0508-5002 https://orcid.org/0000-0001-9672-2064 https://orcid.org/0000-0002-6511-3431 http://dx.doi.org/10.1021/ACSCHEMBIO.5B01083 ACS Chemical Biology Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf American Chemical Society (ACS) PMC |
| spellingShingle | DiChiara, Andrew Stephen Taylor, Rebecca J. Wong, Madeline Y. Doan, Ngoc Duc Del Rosario, Amanda M Shoulders, Matthew D. Mapping and Exploring the Collagen-I Proteostasis Network |
| title | Mapping and Exploring the Collagen-I Proteostasis Network |
| title_full | Mapping and Exploring the Collagen-I Proteostasis Network |
| title_fullStr | Mapping and Exploring the Collagen-I Proteostasis Network |
| title_full_unstemmed | Mapping and Exploring the Collagen-I Proteostasis Network |
| title_short | Mapping and Exploring the Collagen-I Proteostasis Network |
| title_sort | mapping and exploring the collagen i proteostasis network |
| url | http://hdl.handle.net/1721.1/113577 https://orcid.org/0000-0002-9438-0643 https://orcid.org/0000-0002-0508-5002 https://orcid.org/0000-0001-9672-2064 https://orcid.org/0000-0002-6511-3431 |
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