Yifei Yang (Presenter)
University of Chicago
Bio: Dr. Yifei Yang recently defended her PhD thesis, focused on studying the molecular mechanism of PC2 function regulation, in the Department of Pharmacology at Yale University. She is now a Clinical Chemistry Fellow at University of Chicago Medical Center, applying chemical biology tools in clinical diagnostics.
Authorship: Yifei Yang(1),(2), Michael E. Hodsdon (1), Elias J. Lolis (2) and Barbara E. Ehrlich (2),(3)
Departments of (1) Pharmacology, (2) Laboratory Medicine, (3) Cellular and Molecular Physiology, Yale University
| Short Abstract The C-terminal tail of polycystin-2 (PC2 Cterm) is crucial for channel regulation. Mutations in PC2 Cterm can cause autosomal dominant polycystic kidney disease. PC2 forms a calcium-permeable channel in the membrane and its function is regulated by cytosolic calcium-levels. To gain insight into how calcium-binding regulates the channel via its C-terminal tail, we characterized the conformational and dynamic responses of calcium within the PC2 Cterm, using hydrogen-deuterium exchange mass spectrometry. Our study, for the first time, provides a complete map of dynamic responses to calcium-binding within the full length C-terminal tail. Our results suggest mechanisms for functional regulation of the PC2 channel and the roles of PC2 in pathophysiology of polycystic kidney disease. |
Long Abstract
Polycystin-2 (PC2) forms a calcium-permeable channel in the membrane and its function is regulated by cytosolic calcium-levels. Mutations in the C-terminal tail of human PC2 (HPC2 Cterm) lead to autosomal dominant polycystic kidney disease. The HPC2 Cterm protein contains a calclium-binding site responsible for channel gating and function.
To gain insight into how calcium-binding regulates the channel via its C-terminal tail, we characterized calcium-binding and its conformational and dynamic responses within the HPC2 Cterm. By examining hydrogen-deuterium (H-D) exchange profiles, we found that the coiled-coil domain in HPC2 Cterm formed a stable helix bundle regardless of calcium-binding. The HPC2 L1EF construct contains the calcium-binding EF-hand and the N-terminal linker region without the downstream coiled-coil. We found that the N-terminal linker stabilized the calcium-bound conformation of the EF-hand, thus enhancing its calcium-binding affinity to the same level as the full length Cterm. By comparing the conformational dynamics in HPC2 Cterm and L1EF when calcium-bound, we have determined that the coiled-coil domain is not involved in stabilizing the calcium-bound conformation of the EF-hand. Although we found that HPC2 Cterm and L1EF share a similar level of structural stability upon calcium-binding, they have different profiles of H-D exchange dynamics under non-saturating calcium conditions.
Our study, for the first time, provides a complete map of dynamic responses to calcium-binding within the full length C-terminal tail. Our results suggest mechanisms for functional regulation of the PC2 channel and the roles of PC2 in pathophysiology of polycystic kidney disease.
References & Acknowledgements:
References
[1] Gabow PA. Autosomal dominant polycystic kidney disease. N Engl J Med. 1993;329:332-42.
[2] Cai Y, Anyatonwu G, Okuhara D, Lee KB, Yu Z, Onoe T, et al. Calcium dependence of polycystin-2 channel activity is modulated by phosphorylation at Ser812. J Biol Chem. 2004;279:19987-95.
[3] Y. Yang, M.E.Hodsdon, E.J.Lolis, B.E.Ehrlich. Conformational dynamics of Ca2+-dependent responses in the polycystin-2 C-terminal tail. Biochem. J. 2015, 10.1042/BJ20151031
[4] Yang Y, Keeler C, Kuo IY, Lolis EJ, Ehrlich BE, Hodsdon ME. Oligomerization of the polycystin-2 C-terminal tail and effects on its Ca2+-binding properties. J Biol Chem. 2015;290:10544-54.
[5] Celic AS, Petri ET, Benbow J, Hodsdon ME, Ehrlich BE, Boggon TJ. Calcium-induced conformational changes in C-terminal tail of polycystin-2 are necessary for channel gating. J Biol Chem. 2012;287:17232-40.
[6] Keeler C, Poon G, Kuo IY, Ehrlich BE, Hodsdon ME. An explicit formulation approach for the analysis of calcium binding to EF-hand proteins using isothermal titration calorimetry. Biophys J. 2013;105:2843-53.
Acknowledgements
This work was funded by NIH Award No. R21RR032351 to M.E.H. and NIH grants R01 DK087844 to B.E.E. and M.E.H. Y.Y was supported by a pre-doctoral fellowship from CSC-Yale world scholar fellowship.
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