SYNAPTIC FUNCTIONS OF SPECTRIN

血影蛋白的突触功能

• 批准号: 6163121
• 负责人: R K SIHAG
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF NEUROLOGICAL DISORDERS AND STROKE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6163121
• 关键词: actins; crosslink; intermolecular interaction; laboratory rat; membrane proteins; nerve /myelin protein; neural cell adhesion molecules; neural transmission; neurotransmitter transport; protein purification; protein structure function; spectrin; synapses; synaptosomes; tubulin

Spectrin, an alpha/beta heterodimer, is a calcium and calmodulin-binding membrane skeleton protein. It is suspected that spectrin is involved in the formation of distinct membrane domains at the synapse and plays a role in the regulation of synaptic functions, such as, neurotransmission and release of neurotransmitters. To identify the presynaptic or postsynaptic density proteins that interact with spectrin in a domain- specific manner, spectrin and associated proteins were extracted under mild conditions from the synaptosomal fractions, negatively stained or sprayed on mica chips in glycerol and analyzed by electron microscopy. Actin filaments and other so far unidentified globular proteins were found to be associated with the 200 nm spectrin tetramers or oligomers. When the lysed hippocampal neurons or synaptosomal fractions from rat brain were treated with heterobifunctional sulfhydral cross-linking reagents, analyses of the Triton X-100, SDS or SDS and mercaptoethanol soluble fractions showed that spectrin, and the proteins tentatively identified as actin, tubulin, N-CAM and several unidentified membrane proteins were cross-linked although direct cross-linking of any of these proteins with spectrin has yet to be demonstrated. When the hippocampal neuron primary cultures were analyzed by using double immunofluorescence techniques with 4 different anti-spectrin and anti-syntaxin or SNAP-25 antibodies, spectrin was localized underneath the plasma membrane and at the synaptic densities, but spectrin did not show colocalization with the presynaptic syntaxin or SNAP-25 suggesting that these proteins are not directly associated with spectrin. Identification of the proteins with which spectrin directly associates at the synapse will help to understand the role of spectrin in synaptic functions.

Spectrin是一种α/β异源二聚体，是一种钙和钙调蛋白结合蛋白。 膜骨架蛋白怀疑血影蛋白参与了 在突触处形成不同的膜结构域， 在调节突触功能（如神经传递）中的作用 和神经递质的释放为了识别突触前或 与血影蛋白相互作用的突触后密度蛋白 特异性地，在一定条件下提取血影蛋白和相关蛋白。 来自突触体部分的轻度条件，负染色或 喷洒在甘油中的云母片上并通过电子显微镜分析。 肌动蛋白丝和其他迄今尚未鉴定的球状蛋白质， 发现与200 nm血影蛋白四聚体或寡聚体相关。 当大鼠海马神经元或突触体部分裂解后， 脑用异双功能巯基交联处理 Triton X-100、SDS或SDS和巯基乙醇的分析试剂 可溶性组分显示血影蛋白，并且蛋白质初步 鉴定为肌动蛋白、微管蛋白、N-CAM和几种未鉴定的膜 蛋白质是交联的，尽管这些蛋白质中的任何一种的直接交联 具有血影蛋白的蛋白质尚未被证实。当海马 用免疫荧光双标记法分析神经元原代培养物 使用4种不同的抗血影蛋白和抗突触融合蛋白或SNAP-25的技术 抗体，血影蛋白位于质膜下， 在突触密度，但血影蛋白没有显示共定位与 突触前突触融合蛋白或SNAP-25表明这些蛋白质是 与血影蛋白没有直接联系。蛋白质鉴定 血影蛋白在突触处的直接联系将有助于 理解血影蛋白在突触功能中的作用。