scholarly journals Cardiac ryanodine receptor N-terminal region biosensors identify novel inhibitors via FRET-based high-throughput screening

2021 ◽  
pp. 101412
Author(s):  
Jingyan Zhang ◽  
Daniel P. Singh ◽  
Christopher Y. Ko ◽  
Roman Nikolaienko ◽  
Siobhan M. Wong King Yuen ◽  
...  
2020 ◽  
Vol 118 (3) ◽  
pp. 173a-174a
Author(s):  
Jingyan Zhang ◽  
Robyn T. Rebbeck ◽  
David D. Thomas ◽  
Filip V. Petegem ◽  
Razvan L. Cornea

2013 ◽  
Vol 20 (11) ◽  
pp. 1211-1216 ◽  
Author(s):  
L’ubomír Borko ◽  
Július Kostan ◽  
Alexandra Zahradníkova ◽  
Vladimír Pevala ◽  
Juraj Gasperík ◽  
...  

2018 ◽  
Vol 94 (1) ◽  
pp. 722-730 ◽  
Author(s):  
Takashi Murayama ◽  
Nagomi Kurebayashi ◽  
Mari Ishigami-Yuasa ◽  
Shuichi Mori ◽  
Yukina Suzuki ◽  
...  

2020 ◽  
Vol 118 (3) ◽  
pp. 272a
Author(s):  
Manuel Paina ◽  
Jim Goodchild ◽  
Lucy Firth ◽  
Katharina Montag ◽  
Maria Grazia Garibaldi ◽  
...  

2018 ◽  
Vol 114 (3) ◽  
pp. 117a
Author(s):  
Claire E. Haskin ◽  
Robyn T. Rebbeck ◽  
Gregory D. Gillispie ◽  
David D. Thomas ◽  
Razvan L. Cornea

2014 ◽  
Vol 106 (2) ◽  
pp. 108a
Author(s):  
Lynn Kimlicka ◽  
Ching-Chieh Tung ◽  
Anna-Carin C. Carlsson ◽  
Paolo A. Lobo ◽  
Zhiguang Yuchi ◽  
...  

Structure ◽  
2013 ◽  
Vol 21 (8) ◽  
pp. 1440-1449 ◽  
Author(s):  
Lynn Kimlicka ◽  
Ching-Chieh Tung ◽  
Anna-Carin Cecilia Carlsson ◽  
Paolo Antonio Lobo ◽  
Zhiguang Yuchi ◽  
...  

2021 ◽  
Author(s):  
Jingyan Zhang ◽  
Siobhan M. Wong King Yuen ◽  
Jacob A. Schwarz ◽  
Levy M. Treinen ◽  
Ching-Chieh Tung ◽  
...  

AbstractThe N-terminal region (NTR) of the ryanodine receptor (RyR) calcium channels is critical to the regulation of Ca2+ release during excitation-contraction coupling. NTR hosts numerous mutations linked to skeletal and cardiac myopathies (RyR1 and RyR2, respectively), highlighting its potential as therapeutic target. Here, we labeled the NTR of mouse RyR2 at subdomains A, B, and C with donor and acceptor pairs for fluorescence resonance energy transfer (FRET), obtaining two biosensors. Using fluorescence lifetime (FLT)-detection of intramolecular FRET, we developed high-throughput screening (HTS) assays with the biosensors to identify small-molecule modulators of RyR. We screened a 1280-compound validation library and identified several hits. Hits with saturable FRET dose-response profiles, and previously unreported effects on RyR activity, were further tested using [3H]ryanodine binding to isolated sarcoplasmic reticulum vesicles, to measure their effects on full-length RyR opening in its natural membrane environment. We identified three novel inhibitors of both RyR1 and RyR2, and two RyR1-selective inhibitors at nanomolar Ca2+. These compounds may function as inhibitors of leaky RyRs in muscle. Two of these hits activated RyR1 only at micromolar Ca2+, highlighting them as potential activators of excitation-contraction coupling. These results indicate that large-scale HTS using this platform can lead to compounds with potential for therapeutic development.


2016 ◽  
Vol 22 (2) ◽  
pp. 176-186 ◽  
Author(s):  
Robyn T. Rebbeck ◽  
Maram M. Essawy ◽  
Florentin R. Nitu ◽  
Benjamin D. Grant ◽  
Gregory D. Gillispie ◽  
...  

Using time-resolved fluorescence resonance energy transfer (FRET), we have developed and validated the first high-throughput screening (HTS) method to discover compounds that modulate an intracellular Ca2+ channel, the ryanodine receptor (RyR), for therapeutic applications. Intracellular Ca2+ regulation is critical for striated muscle function, and RyR is a central player. At resting [Ca2+], an increased propensity of channel opening due to RyR dysregulation is associated with severe cardiac and skeletal myopathies, diabetes, and neurological disorders. This leaky state of the RyR is an attractive target for pharmacological agents to treat such pathologies. Our FRET-based HTS detects RyR binding of accessory proteins calmodulin (CaM) or FKBP12.6. Under conditions that mimic a pathological state, we carried out a screen of the 727-compound NIH Clinical Collection, which yielded six compounds that reproducibly changed FRET by >3 SD. Dose–response of FRET and [3H]ryanodine binding readouts reveal that five hits reproducibly alter RyR1 structure and activity. One compound increased FRET and inhibited RyR1, which was only significant at nM [Ca2+], and accentuated without CaM present. These properties characterize a compound that could mitigate RyR1 leak. An excellent Z′ factor and the tight correlation between structural and functional readouts validate this first HTS method to identify RyR modulators.


Sign in / Sign up

Export Citation Format

Share Document