We propose to elucidate the molecular mechanisms of arrestin-1 interactions with different functional forms of rhodopsin, which significantly contribute to exquisitely regulated and precisely timed function of photoreceptor cells, using the combination of novel cutting-edge biophysical methods and in vivo experiments. We will compare the shape of the complex of arrestin-1 with two forms of phospho-rhodopsin: rhodopsin carrying 3-4 phosphates, which are routinely formed in rod photoreceptors and appears to be perfectly harmless, and tight arrestin-1 complexes with hyper-phosphorylated rhodopsin that are cytotoxic in mice and Drosophila. We will explore the dynamics of free and rhodopsin-bound arrestin using high pressure EPR to elucidate the functional role of its flexibility. We will determine the biological role of arrestin-1 self-association in photoreceptors in vivo, which is currently unknown. Finally, we will test novel compensational approach to gene therapy of gain-of-function rhodopsin mutations using engineered enhanced arrestin-1 mutants capable of shutting off rhodopsin signaling independently of its phosphorylation. Proposed studies will significantly improve our understanding of photoreceptor physiology and advance our progress towards gene therapy of gain-of-function rhodopsin mutations. Due to high conservation of mechanics of GPCR regulation, mechanistic studies of arrestin-1 binding to rhodopsin will have broader implications, improving our understanding of the molecular basis of the function of other arrestin subtypes. Arrestin will be the first signaling protein studied by high-pressure EPR, which will reveal the functional role of its conformational flexibility. Protein-protein interactions, whch are often mediated by flexible or even unstructured elements, play key role in the regulation of cellular processes. Therefore, proposed studies will shed new light on many aspects of cell signaling. Our strategic goal is to gain sufficient understanding of protein-protein interactions that govern cell signaling to construct mutants with desired functional characteristics for research and therapeutic purposes.
We propose to elucidate the molecular mechanisms of arrestin-1 interactions with different functional forms of rhodopsin, which significantly contribut to exquisitely regulated and precisely timed function of photoreceptor cells. To this end, we propose to use the combination of novel cutting-edge biophysical methods and in vivo experiments. We also propose to test novel compensational approach to gene therapy of gain-of-function rhodopsin mutations using engineered enhanced arrestin-1 mutants capable of shutting off rhodopsin signaling independently of its phosphorylation.
| Gurevich, Vsevolod V; Gurevich, Eugenia V; Uversky, Vladimir N (2018) Arrestins: structural disorder creates rich functionality. Protein Cell 9:986-1003 |
| Chen, Qiuyan; Iverson, Tina M; Gurevich, Vsevolod V (2018) Structural Basis of Arrestin-Dependent Signal Transduction. Trends Biochem Sci 43:412-423 |
| Tso, Shih-Chia; Chen, Qiuyan; Vishnivetskiy, Sergey A et al. (2018) Using two-site binding models to analyze microscale thermophoresis data. Anal Biochem 540-541:64-75 |
| Vishnivetskiy, Sergey A; Sullivan, Lori S; Bowne, Sara J et al. (2018) Molecular Defects of the Disease-Causing Human Arrestin-1 C147F Mutant. Invest Ophthalmol Vis Sci 59:13-20 |
| Gurevich, Vsevolod V; Gurevich, Eugenia V (2018) GPCRs and Signal Transducers: Interaction Stoichiometry. Trends Pharmacol Sci 39:672-684 |
| Cleghorn, Whitney M; Bulus, Nada; Kook, Seunghyi et al. (2018) Non-visual arrestins regulate the focal adhesion formation via small GTPases RhoA and Rac1 independently of GPCRs. Cell Signal 42:259-269 |
| Chen, Qiuyan; Perry, Nicole A; Vishnivetskiy, Sergey A et al. (2017) Structural basis of arrestin-3 activation and signaling. Nat Commun 8:1427 |
| Indrischek, Henrike; Prohaska, Sonja J; Gurevich, Vsevolod V et al. (2017) Uncovering missing pieces: duplication and deletion history of arrestins in deuterostomes. BMC Evol Biol 17:163 |
| Zhu, Lu; Rossi, Mario; Cui, Yinghong et al. (2017) Hepatic ?-arrestin 2 is essential for maintaining euglycemia. J Clin Invest 127:2941-2945 |
| Gurevich, Vsevolod V; Gurevich, Eugenia V (2017) Molecular Mechanisms of GPCR Signaling: A Structural Perspective. Int J Mol Sci 18: |
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