Regulation of Kv2 potassium channel diversity and function in brain by electrically silent subunits

电沉默亚基对大脑中 Kv2 钾通道多样性和功能的调节

• 批准号: 10288261
• 负责人: MICHAEL J FERNS
• 金额: $ 43.18万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10288261
• 关键词: Action Potentials; Address; Antibodies; Behavior; Binding; Brain; Brain region; C-terminal; Cell membrane; Cells; DNA Sequence Alteration; Dendrites; Endoplasmic Reticulum; Epilepsy; Exhibits; Family; Family member; Functional disorder; Gene Expression Profiling; Genes; Homeostasis; Homo; Hyperactivity; Hypoxia; Immunoblotting; Ion Channel; Ions; Knock-out; Knowledge; Kv2.1 channel; Label; Lead; Lipids; Mass Spectrum Analysis; Membrane; Mutation; Neurons; Pattern; Phosphorylation; Play; Population; Potassium Channel; Proteins; Proteomics; Regulation; Rodent; Role; Sampling; Seizures; Signal Transduction; Site; Source; Stimulus; Structure; Tail; Testing; Time; Voltage-Gated Potassium Channel; Work; base; biophysical properties; crosslink; defined contribution; differential expression; experimental study; hippocampal pyramidal neuron; insight; mRNA Expression; nervous system disorder; neuronal cell body; response; selective expression; voltage

Voltage-gated K+ channels of the Kv2 family (Kv2.1 and 2.2) are highly expressed in brain and have important and diverse functions in mammalian neurons. First, Kv2 channels play important canonical roles as voltage-activated channels that regulate neuronal action potentials and membrane excitability, and genetic mutations or targeted deletions of these channels cause hyperactivity and seizures. Second, Kv2 channels are highly clustered on neuronal cell bodies and proximal dendrites and have a separate structural role in organizing specialized microdomains called endoplasmic reticulum - plasma membrane (ER-PM) junctions, which serve as important hubs for Ca and lipid signaling in brain neurons. Kv2 channels organize these sites by interacting with ER VAP proteins via a highly conserved PRC domain in the Kv2 C-terminal tail. Kv2 channels are thought to be largely composed of Kv2.1 and Kv2.2 subunits, which can co-assemble to form homo- or hetero-tetrameric channels. Another potential - but largely unexplored - source of Kv2 channel diversity comes from electrically silent or accessory K+ channel (KvS) subunit genes. This extensive and highly conserved family was termed “electrically silent” because they require co-assembly together with Kv2.1 or Kv2.2 in order to form functional ion-conducting channels when expressed in heterologous cells. Gene expression analyses demonstrate that KvS subunit family members are differentially expressed in distinct brain regions and neuron subtypes, however there is currently no information available on native KvS-Kv2 channels in brain at the protein level. In preliminary mass spectrometry-based proteomics experiments, we identified several KvS subunits that interact with Kv2.1 in brain and generated specific antibodies to these subunits. Using these antibodies, we have made the major discovery that one KvS subunit is a remarkably common constituent of Kv2 channels and that it colocalizes with Kv2 at ER-PM junctions in subsets of cortical neurons. The objective of this exploratory study, therefore, is to characterize the neuronal expression and subcellular localization of these KvS family members, and to define their contribution to Kv2 channel diversity in mammalian brain neurons. This exploratory work will provide important new insight into how selective expression of KvS subunits impacts Kv2 channel diversity in brain and how it modulates Kv2 channel function and localization in different neuronal populations.

Kv 2家族的电压门控K+通道（Kv2.1和2.2）在脑中高度表达，并且具有重要的生物学功能。 以及哺乳动物神经元的多种功能。首先，Kv 2通道作为调节神经元动作电位和膜兴奋性的电压激活通道发挥重要的典型作用，并且基因突变或 这些通道的靶向缺失导致多动症和癫痫发作。第二，Kv 2通道高度 聚集在神经元细胞体和近端树突上，在组织中具有单独的结构作用 专门的微结构域称为内质网-质膜（ER-PM）连接， 大脑神经元中钙和脂质信号的重要枢纽。Kv 2通道通过与 ER VAP蛋白通过Kv 2 C-末端尾中高度保守的PRC结构域。Kv 2通道被认为是 主要由Kv2.1和Kv2.2亚基组成，它们可以共组装形成同源或异源四聚体 渠道Kv 2通道多样性的另一个潜在的（但在很大程度上未被探索的）来源来自电 沉默或辅助K+通道（KvS）亚基基因。这个广泛而高度保守的家族被称为 “电沉默”，因为它们需要与Kv2.1或Kv2.2共同组装在一起，以便形成功能性的 当在异源细胞中表达时，基因表达分析表明， 然而，KvS亚基家族成员在不同的大脑区域和神经元亚型中差异表达， 目前还没有关于脑中天然KvS-Kv 2通道在蛋白质水平上的信息。初步 基于质谱的蛋白质组学实验，我们确定了几个与Kv2.1相互作用的KvS亚基。 并产生针对这些亚单位的特异性抗体。利用这些抗体，我们已经使主要的 发现一个KvS亚基是Kv 2通道的一个非常常见的组成部分，并且它与 皮质神经元亚群中ER-PM连接处的Kv 2。因此，本探索性研究的目的是 表征这些KvS家族成员的神经元表达和亚细胞定位，并定义 它们对哺乳动物脑神经元中Kv 2通道多样性的贡献。这项探索性工作将提供 关于KvS亚基的选择性表达如何影响脑中Kv 2通道多样性的重要新见解， 它如何调节Kv 2通道功能和定位在不同的神经元群体。

项目成果

Electrically silent KvS subunits associate with native Kv2 channels in brain and impact diverse properties of channel function.

电沉默 KvS 亚基与大脑中的天然 Kv2 通道相关联，并影响通道功能的多种特性。

• DOI: 10.1101/2024.01.25.577135
• 发表时间: 2024
• 期刊: bioRxiv : the preprint server for biology
• 作者: Ferns,Michael;vanderList,Deborah;Vierra,NicholasC;Lacey,Taylor;Murray,Karl;Kirmiz,Michael;Stewart,RobertG;Sack,JonT;Trimmer,JamesS
• 通讯作者: Trimmer,JamesS