Abstract
Collagen overproduction is a feature of fibrosis and cancer, while insufficient deposition of functional collagen molecules and/or the secretion of malformed collagen is common in genetic disorders like osteogenesis imperfecta. Collagen secretion is an appealing therapeutic target in these and other diseases, as secretion directly connects intracellular biosynthesis to collagen deposition and biological function in the extracellular matrix. However, small molecule and biological methods to tune collagen secretion are severely lacking. Their discovery could prove useful not only in the treatment of disease, but also in providing tools for better elucidating mechanisms of collagen biosynthesis. We developed a cell-based, high-throughput luminescent assay of collagen type I secretion and used it to screen for small molecules that selectively enhance or inhibit that process. Among several validated hits, the Hsp90 inhibitor 17-allylaminogeldanamycin (17-AAG) robustly decreases the secretion of collagen-I by our model cell line and by human primary cells. In these systems, 17-AAG and other pan-isoform Hsp90 inhibitors reduce collagen-I secretion post-translationally and are not global inhibitors of protein secretion. Surprisingly, the consequences of Hsp90 inhibitors cannot be attributed to inhibition of the endoplasmic reticulum's Hsp90 isoform, Grp94. Instead, collagen-I secretion likely depends on the activity of cytosolic Hsp90 chaperones, even though such chaperones cannot directly engage nascent collagen molecules. Our results highlight the value of a cell-based high-throughput screen for selective modulators of collagen secretion and suggest an unanticipated role for cytosolic Hsp90 in collagen secretion.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 2814-2827 |
| Number of pages | 14 |
| Journal | Biochemistry |
| Volume | 57 |
| Issue number | 19 |
| DOIs | |
| State | Published - May 15 2018 |
| Externally published | Yes |
Bibliographical note
Funding Information:This work was supported by the National Institutes of Health (Grants 1R03AR067503 and 1R01AR071443), the 56th Edward Mallinckrodt Jr. Foundation Faculty Scholar Award, and the Massachusetts Institute of Technology Department of Chemistry (to M.D.S.). This work was also supported in part by an endowment from the Roger and Dorothy Hirl Research Fund and a Career Development Award, as well as an unrestricted grant from Research to Prevent Blindness (to J.D.H.). M.Y.W. was supported by a National Science Foundation graduate research fellowship and a Prof. Amar G. Bose Research Grant. N.D.D. was supported by a Canadian Institutes of Health Research (CIHR) postdoctoral fellowship. A.S.D. was supported by a National Institutes of Health Ruth L. Kirschstein predoctoral fellowship (1F31AR067615). L.J.P. was supported by a National Science Foundation graduate research fellowship. Additional funding was provided by a National Cancer Institute core grant (P30-CA14051) to the Koch Institute and an NEI Visual Science Core Grant (EY020799) to the University of Texas Southwestern Medical Center.
Funding Information:
*E-mail: [email protected]. Telephone: (617) 452-3525. Fax: (617) 369-7500. ORCID Matthew D. Shoulders: 0000-0002-6511-3431 Author Contributions M.Y.W. and M.D.S. conceived the project. M.Y.W., M.D.S., and J.D.H. designed experiments and analyzed data. M.Y.W., N.D.D., A.S.D., L.J.P., J.H.C., C.K.S., and N.W. generated reagents and performed experiments. M.Y.W. and M.D.S. drafted the paper. All authors edited the manuscript. Funding This work was supported by the National Institutes of Health (Grants 1R03AR067503 and 1R01AR071443), the 56th Edward Mallinckrodt Jr. Foundation Faculty Scholar Award, and the Massachusetts Institute of Technology Department of Chemistry (to M.D.S.). This work was also supported in part by an endowment from the Roger and Dorothy Hirl Research Fund and a Career Development Award, as well as an unrestricted grant from Research to Prevent Blindness (to J.D.H.). M.Y.W. was supported by a National Science Foundation graduate research fellowship and a Prof. Amar G. Bose Research Grant. N.D.D. was supported by a Canadian Institutes of Health Research (CIHR) postdoctoral fellowship. A.S.D. was supported by a National Institutes of Health Ruth L. Kirschstein predoctoral fellowship (1F31AR067615). L.J.P. was supported by a National Science Foundation graduate research fellowship. Additional funding was provided by a National Cancer Institute core grant (P30-CA14051) to the Koch Institute and an NEI Visual Science Core Grant (EY020799) to the University of Texas Southwestern Medical Center. Notes The authors declare no competing financial interest.
Publisher Copyright:
© 2018 American Chemical Society.
