Skeletal Muscle Ryanodine Receptor Permeation and Self Counter-Ion Flow

骨骼肌 Ryanodine 受体渗透和自反离子流

• 批准号: 7488500
• 负责人: Michael Fill
• 金额: $ 31.18万
• 依托单位: RUSH UNIVERSITY MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7488500
• 关键词: Address; Affect; Affinity; Amino Acid Motifs; Amino Acids; Amitriptyline; Antibodies; Appendix; Arts; Attenuated; Binding; Biological Assay; Biological Models; Budgets; Caffeine; Caliber; Calmodulin; Calsequestrin; Cations; Cells; Central Core Myopathy; Characteristics; Charge; Condition; Coupled; Data; Defect; Detection; Dimensions; Disadvantaged; Disease; Disputes; Dose; Electric Capacitance; Electrodes; Event; Experimental Designs; Figs - dietary; Flavoring; Free Energy; Frequencies; Hand; Helix (Snails); Image; Immunoblotting; Ion Channel; Ions; Knowledge; Left; Life; Light; Lipid Bilayers; Localized; Location; Measurement; Measures; Mediating; Mediator of activation protein; Membrane; Membrane Potentials; Methods; Microsomes; Modeling; Molecular; Mus; Muscle; Mutation; Nature; Noise; Operative Surgical Procedures; Oral cavity; Outcome; Pathway interactions; Pharmaceutical Preparations; Phase; Physiological; Pilot Projects; Play; Positioning Attribute; Postdoctoral Fellow; Potassium Channel; Preparation; Principal Investigator; Probability; Process; Property; Proteins; Protocols documentation; Publications; Published Comment; Publishing; Rate; Rattus; Regulation; Relative (related person); Research; Research Personnel; Research Support; Resolution; Risk; Role; RyR1; RyR2; Ryanodine; Ryanodine Receptor Calcium Release Channel; Sarcoplasmic Reticulum; Score; Series; Side; Signal Transduction; Site; Skeletal Muscle; Skeletal system; Structure; Study Section; Suggestion; Tacrolimus Binding Protein 1A; Techniques; Teflon; Testing; Therapeutic Intervention; Thinking; Time; Trees; Triflupromazine; Vesicle; Work; attenuation; base; cell type; computerized data processing; concept; driving force; ear helix; experience; forest; inositol-1,4,5-triphosphate receptor; mouse model; mutant; novel; programs; protein expression; reconstitution; research study; response; size; theories; uptake; voltage; voltage clamp

DESCRIPTION (provided by applicant): 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.

描述（由申请人提供）：骨骼肌的去极化通过肌浆网（SR）中的 1 型兰尼碱受体（RyR1）Ca 释放通道启动 Ca 释放。在离散的 SR Ca 释放位点协调打开多个 RyR1 通道会产生称为 Ca 火花的小型局部释放事件，这是细胞中 SR Ca 释放的基本单位。火花募集/求和是产生驱动肌肉收缩性的全局 Ca 释放瞬态的原因。许多研究都集中在 Ca 释放瞬态的开始和终止。很少有人关注 RyR 传导/选择性，而且（据我们所知）没有人研究 RyR1 的 RyR 渗透特性对细胞 Ca 信号传导现象（如 Ca 释放瞬态）的生理影响。 RyR 孔被认为具有类似于细菌 K 通道 (KcsA) 的结构。 RyR 具有一个腔环，其中包含预测的孔螺旋和被识别为选择性过滤器的氨基酸基序 (GGGIG)。与 KcsA 孔不同，RyR 孔电导率高，选择性差。已发表的作品以及我们自己的初步数据揭示了一些关键的分子决定因素，这些决定因素定义了 RyR1 孔的高电导率和差选择性的特征。这包括一些与中央核心疾病（CC​​D）相关的自然发生的突变体。该提案结合这些信息来定义生理条件下控制 RyR1 渗透的机制，并应用这些知识来定义单通道 RyR1 渗透过程的一些关键生理后果。检验以下假设。假设：在细胞中，RyR1 通道的选择性较差，导致多种离子（Ca、Mg 和 K）竞争开放孔的占据。虽然这种竞争减弱了通过开孔的净 SR Ca 流出，但它也允许孔在 Ca 释放过程中调节其自身的反离子通量。这种自抗衡离子流有效地将局部 SR 膜电势限制在远离 Ca 能斯特电势 (ECa) 的位置，使得反式 SR Ca 驱动力主要取决于反式 SR Ca 浓度梯度。这在生理上很重要，因为它允许 RyR 通道在较长时间内（> 5 ms 或 Ca 瞬变的上升时间）维持 Ca 释放。具体目标是： (1) 定义 RyR1 渗透的已知分子决定因素如何组合以产生/影响开放 RyR1 孔的电导率和选择性。 (2) 定义RyR1孔选择性相对较差的功能后果；即通过开孔的净 SR Ca 流出量的衰减以及开孔在 SR Ca 流出期间携带其自身反离子通量的能力。