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: 7683996
• 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/7683996
• 关键词: Amino Acid Motifs; Amino Acids; Attenuated; Biological Assay; Cations; Cells; Central Core Myopathy; Characteristics; Charge; Data; Detection; Dimensions; Disadvantaged; Disease; Event; Image; Ions; Knowledge; Lipid Bilayers; Location; Mediating; Membrane Potentials; Modeling; Molecular; Muscle; Mutation; Physiological; Potassium Channel; Process; Publishing; RyR1; Ryanodine Receptor Calcium Release Channel; Ryanodine Receptors; Sarcoplasmic Reticulum; Signal Transduction; Site; Skeletal Muscle; Structure; Testing; Time; Vesicle; Work; attenuation; base; cell type; computerized data processing; driving force; mutant; novel; theories

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(申请人提供)：骨骼肌去极化通过肌浆网(SR)中的1型Ryanodine受体(RyR1)钙释放通道启动钙释放。在离散的SR钙释放部位协同开放多个RyR1通道会产生称为钙火花的小的局部性释放事件，这是细胞内SR钙释放的基本单位。火花募集/总和是产生推动肌肉收缩的全球钙释放瞬变的原因。许多研究都集中在钙释放瞬变的开始和终止。很少有人关注RyR的传导/选择性，还没有人(据我们所知)研究RyR的渗透特性对细胞内钙信号的影响，如钙释放瞬变。RyR孔被认为具有类似于细菌K通道(KCSA)的结构。RyR有一个管腔环，其中包含一个预测的毛孔螺旋和一个被鉴定为选择性过滤器的氨基酸基序(GGGIG)。与KCSA孔不同，RyR孔具有高电导和较差的选择性。已发表的工作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、低选择性的特点。这包括一些与中枢性核心病相关的自然发生的突变。本建议结合这些信息来定义在生理条件下控制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分支。对以下假设进行了检验。假设：在细胞中，RyR1通道的低选择性允许多个离子(钙、镁和钾)竞争开放的孔道。虽然这种竞争减弱了通过开放的毛孔的净SR钙外流，但它也允许毛孔在钙释放过程中调节自己的反离子流量。这种自反离子流有效地钳制了远离钙能斯特电位(ECA)的局部SR膜电位，使反式SR钙驱动力主要依赖于反式SR钙浓度梯度。这在生理上很重要，因为它允许RyR通道在较长的一段时间内(&gt；5ms或瞬时钙上升时间)维持钙释放。具体目的是：(1)确定RyR1渗透的已知分子决定因素如何结合以产生/影响开放的RyR1孔的电导和选择性。(2)确定RyR1孔相对较差的选择性的功能后果；即通过开口孔的净SR Ca外流的衰减以及在SR Ca外流过程中开口孔携带其自身反离子通量的能力。