Depolarization of skeletal muscle initiates Ca release through type-1 ryanodine receptor (RyR1) Ca release channels in the sarcoplasmic reticulum (SR). Concerted opening of multiple RyR1 channels at discrete SR Ca release sites generates small localized release events called Ca sparks, the elemental unit of SR Ca release in cells. Spark recruitment/summation is what generates the global Ca release transient that drives muscle contractility. Many studies have focused on the onset and termination of the Ca release transient. Fewer have focused on the RyR conduction/selectivity and none (to our knowledge) have studied what physiological ramifications the RyR permeation characteristics of RyR1 have on cellular Ca signaling phenomena like the Ca release transient. The RyR pore is thought to have a structure analogous to that of the bacterial K channel (KcsA). The RyR has a lumenal loop which contains a predicted pore helix and an amino acid motif (GGGIG) identified as a selectivity filter. Unlike the KcsA pore, the RyR pore has high conductance and is poorly selective. Published works as well as our own preliminary data have revealed some key molecular determinants that define the characteristic high conductance, poor selectivity of the RyR1 pore. This includes some naturally occurring mutants associated with central core disease (CCD). This proposal combines this information to define the mechanisms that govern RyR1 permeation under physiological conditions and applies this knowledge to define some key physiological ramifications of the single channel RyR1 permeation process. The following hypothesis is tested. Hypothesis: In cells, the poor selectivity of the RyR1 channel allows multiple ions (Ca, Mg &K) to compete for occupancy of the open pore. While this competition attenuates the net SR Ca efflux through the open pore, it also allows the pore to mediate its own counter ion flux during the Ca release process. This self counter-ion flow effectively clamps local SR membrane potential far from the Ca Nernst potential (ECa) making the trans-SR Ca driving force primarily dependent on the trans-SR Ca concentration gradient. This is physiologically important because it allows the RyR channel to sustain Ca release over an extended period of time (>5 ms or the rise time of Ca transient).
The specific aims are: (1) Define how known molecular determinants of RyR1 permeation combine to generate/influence the conductance and selectivity of the open RyR1 pore. (2) Define functional consequences of the relatively poor selectivity of the RyR1 pore;namely attenuation of net SR Ca efflux through the open pore and the capacity of the open pore to carry its own counter ion flux during SR Ca efflux.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR054098-03
Application #
7683996
Study Section
Skeletal Muscle and Exercise Physiology Study Section (SMEP)
Program Officer
Boyce, Amanda T
Project Start
2007-09-01
Project End
2011-08-31
Budget Start
2009-09-01
Budget End
2010-08-31
Support Year
3
Fiscal Year
2009
Total Cost
$311,836
Indirect Cost
Name
Rush University Medical Center
Department
Physiology
Type
Schools of Medicine
DUNS #
068610245
City
Chicago
State
IL
Country
United States
Zip Code
60612
Zsolnay, Vilmos; Fill, Michael; Gillespie, Dirk (2018) Sarcoplasmic Reticulum Ca2+ Release Uses a Cascading Network of Intra-SR and Channel Countercurrents. Biophys J 114:462-473
Yan, Jiajie; Zhao, Weiwei; Thomson, Justin K et al. (2018) Stress Signaling JNK2 Crosstalk With CaMKII Underlies Enhanced Atrial Arrhythmogenesis. Circ Res 122:821-835
Berti, Claudio; Zsolnay, Vilmos; Shannon, Thomas R et al. (2017) Sarcoplasmic reticulum Ca2+, Mg2+, K+, and Cl- concentrations adjust quickly as heart rate changes. J Mol Cell Cardiol 103:31-39
Uehara, Akira; Murayama, Takashi; Yasukochi, Midori et al. (2017) Extensive Ca2+ leak through K4750Q cardiac ryanodine receptors caused by cytosolic and luminal Ca2+ hypersensitivity. J Gen Physiol 149:199-218
Manno, Carlo; Figueroa, Lourdes C; Gillespie, Dirk et al. (2017) Calsequestrin depolymerizes when calcium is depleted in the sarcoplasmic reticulum of working muscle. Proc Natl Acad Sci U S A 114:E638-E647
Ramos-Franco, Josefina; Fill, Michael (2016) Approaching ryanodine receptor therapeutics from the calcin angle. J Gen Physiol 147:369-73
Berti, Claudio; Furini, Simone; Gillespie, Dirk (2016) PACO: PArticle COunting Method To Enforce Concentrations in Dynamic Simulations. J Chem Theory Comput 12:925-9
Bovo, Elisa; Mazurek, Stefan R; Fill, Michael et al. (2015) Cytosolic Ca²? buffering determines the intra-SR Ca²? concentration at which cardiac Ca²? sparks terminate. Cell Calcium 58:246-53
Gillespie, Dirk; Xu, Le; Meissner, Gerhard (2014) Selecting ions by size in a calcium channel: the ryanodine receptor case study. Biophys J 107:2263-73
Guo, Tao; Nani, Alma; Shonts, Stephen et al. (2013) Sarcoplasmic reticulum K(+) (TRIC) channel does not carry essential countercurrent during Ca(2+) release. Biophys J 105:1151-60

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