BRAIN SPECTRIN AND SYNAPTIC TRANSMISSION

脑血影蛋白和突触传递

• 批准号: 6151571
• 负责人: STEVEN Richard GOODMAN
• 金额: $ 23.34万
• 依托单位: UNIVERSITY OF SOUTH ALABAMA
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-02-01 至 2002-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6151571
• 关键词: actins; brain metabolism; calcium; hippocampus; intermolecular interaction; laboratory rat; neural transmission; protein biosynthesis; protein structure function; proteolysis; recombinant proteins; site directed mutagenesis; spectrin; synapses; synaptic vesicles; synthetic peptide; tissue /cell culture; voltage /patch clamp

This is a new application to study the role of brain spectrin in synaptic transmission. We will determine whether spectrin plays a direct role in depolarization-induced release of neurotransmitter from the presynaptic terminal. The specific hypothesis to be tested is that spectrin is an essential docking protein and Ca2+ sensor protein in the Ca2+-regulated vesicle fusion and neurotransmitter release from 50 nm diameter spherical synaptic vesicles. The aims are to (1) Determine the rate of spectrin synthesis, turnover, and assembly in hippocampal neurons in the resting and depolarized state (+/-Ca2+). (2) Determine whether the proposed binding site for synapsin I on brain beta spectrin (res. 207-445 of beta(Sp)IIsigma1) can bind synaptic vesicles, and define the smallest functional subdomain and essential amino acids for attachment. Also determine whether synaptic vesicle binding displaces the attachment of spectrin and f-actin as proposed in the "casting the line" hypothesis. Finally determine whether the interaction of synaptic vesicles with brain beta spectrin is directly regulated by free Ca2+ levels. (3) Determine whether microinjection of beta spectrin recombinant peptides and synthetic peptides which contain the synaptic vesicle binding domain (and FAB fragments against these synthetic peptides) alters neurotransmitter release from paired hippocampal neurons in culture. Peptides and antibodies will be injected into the presynaptic neurons and we will use patch clamp techniques and quantal analysis to study changes in transmitter release from small spherical synaptic vesicles. Dr. Goodman and his collaborators are in a unique position to successfully conduct these studies because they have developed the necessary spectrin DNA constructs, recombinant peptides, synthetic peptides and antibodies to perform the proposed studies. They also have the requisite expertise in neuronal cell culture, molecular biology, protein chemistry, electrophysiology and a long history of working with the spectrin molecule.

这是研究脑血影蛋白的作用的新应用 突触传递。 我们将确定血影蛋白是否发挥作用 去极化诱导的神经递质释放的直接作用 突触前末梢。 待检验的具体假设为 血影蛋白是一种重要的对接蛋白和 Ca2+ 传感器蛋白 Ca2+调节的囊泡融合和神经递质释放 直径 50 nm 的球形突触小泡。 目标是 (1) 确定血影蛋白合成、周转和组装的速率 海马神经元处于静息和去极化状态（+/-Ca2+）。 (2) 确定突触蛋白 I 的拟议结合位点是否在 脑 β 血影蛋白（beta(Sp)IIsigma1 的 res. 207-445）可以结合 突触小泡，并定义最小的功能子域和 附着所必需的氨基酸。还要确定是否有突触 囊泡结合取代了血影蛋白和 f-肌动蛋白的附着 提出了“铸线”假说。最后确定 突触小泡是否与脑β血影蛋白相互作用 直接受游离 Ca2+ 水平调节。 (3)判断是否 β血影蛋白重组肽和合成肽的显微注射 含有突触小泡结合结构域（和 FAB 针对这些合成肽的片段）改变神经递质 从培养物中成对的海马神经元释放。 肽和 抗体将被注射到突触前神经元中，我们将 使用膜片钳技术和定量分析来研究 小球形突触囊泡释放递质。 博士。 古德曼和他的合作者处于独特的地位，能够成功地 进行这些研究是因为他们已经开发出了必要的 血影蛋白 DNA 构建体、重组肽、合成肽和 抗体来进行拟议的研究。他们还拥有 神经细胞培养、分子生物学方面必要的专业知识， 蛋白质化学、电生理学和悠久的工作历史 与血影蛋白分子。

项目成果

Synthesis, assembly, and turnover of alpha and beta-erythroid and nonerythroid spectrins in rat hippocampal neurons.

大鼠海马神经元中α和β-红系血影蛋白和非红系血影蛋白的合成、组装和周转。

• DOI: 10.1016/s0006-8993(99)02026-0
• 发表时间: 1999
• 期刊: Brain research
• 影响因子: 2.9
• 作者: Sangerman,J;Gard,AL;Shah,A;Goodman,SR
• 通讯作者: Goodman,SR

Spectrin ubiquitination and oxidative stress: potential roles in blood and neurological disorders.

血影蛋白泛素化和氧化应激：在血液和神经系统疾病中的潜在作用。

• 发表时间: 2001
• 期刊: Cellular & molecular biology letters.
• 作者: Sangerman,J;Kakhniashvili,D;Brown,A;Shartava,A;Goodman,SR
• 通讯作者: Goodman,SR

Essential control of an endothelial cell ISOC by the spectrin membrane skeleton.

光谱膜骨架对内皮细胞ISOC的基本控制。

• DOI: 10.1083/jcb.200106156
• 发表时间: 2001-09-17
• 期刊: JOURNAL OF CELL BIOLOGY
• 影响因子: 7.8
• 作者: Wu, SW;Sangerman, J;Li, M;Brough, GH;Goodman, SR;Stevens, T
• 通讯作者: Stevens, T

Alpha-spectrins are major ubiquitinated proteins in rat hippocampal neurons and components of ubiquitinated inclusions in neurodegenerative disorders.

α-血影蛋白是大鼠海马神经元中的主要泛素化蛋白，也是神经退行性疾病中泛素化包涵体的成分。

• DOI: 10.1016/s0361-9230(01)00426-9
• 发表时间: 2001
• 期刊: Brain research bulletin
• 影响因子: 3.8
• 作者: Sangerman,J;Killilea,A;Chronister,R;Pappolla,M;Goodman,SR
• 通讯作者: Goodman,SR

The domain of brain beta-spectrin responsible for synaptic vesicle association is essential for synaptic transmission.

负责突触小泡关联的大脑β-血影蛋白域对于突触传递至关重要。

• DOI: 10.1016/s0006-8993(00)02796-7
• 发表时间: 2000
• 期刊: Brain research
• 影响因子: 2.9
• 作者: Zimmer,WE;Zhao,Y;Sikorski,AF;Critz,SD;Sangerman,J;Elferink,LA;Xu,XS;Goodman,SR
• 通讯作者: Goodman,SR