MOLECULAR BASIS OF NON-NMDA GLUTAMATE RECEPTOR ASSEMBLEY

离子型谷氨酸受体介导快速兴奋性神经传递。谷氨酸受体亚基的异源多聚化决定了关键的通道特性，这反过来又会影响细胞信号传导过程。然而，亚基寡聚化仅发生在谷氨酸受体亚家族内，表明决定亚基特异性组装的蛋白质结构域。这些实验的目的是确定确定谷氨酸受体亚基寡聚化的分子机制。该片段还对AMPA电流表达产生显性负效应，但对NMDA电流表达不产生显性负效应。除其他外，这些数据表明AMPA受体的氨基末端参与亚基结合，并且可能对组装至关重要。本文提出的实验旨在通过回答几个具体问题来补充这些结果。具体目标#1通过回答几个具体问题来应用强大的酵母双杂交方法结果。具体目标#1应用强大的酵母双杂交方法来定义和表征亚基缔合的最小区域。具体目标#2回答了糖基化是否影响AMPA受体中亚基缔合的问题。具体目标#3中的分析性超离心实验旨在测定氨基末端片段的分子量、化学计量和缔合性质，其已在共免疫沉淀实验中显示出自缔合。最后，具体目标#4旨在确定AMPA受体组装是否发生在内质网或另一个细胞内区室中。结合以前的数据，这些实验将提供AMPA受体生物发生的全面描述。

Ionotropic glutamate receptors mediate fast excitatory neurotransmission. Heteromultimerization of glutamate receptor subunits determines critical channel properties which in turn can influences cell signaling processes. Subunit oligomerization, however, occurs only within glutamate receptor subfamilies, suggesting a protein domain that determines subunit-specific assembly . The objective of these experiments is to define molecular mechanisms that determine oligomerization of glutamate receptor subunits. This fragment also exerts a dominant negative effect on AMPA, but not NMDA, current expression. These data, among others, demonstrate that the amino terminus of AMPA receptors is involved in subunit association and may be critical for assembly. Experiments proposed herein are designed to complement these results by answering several specific questions. Specific Aim #1 applies the powerful yeast- two-hybrid approach results by answering several specific questions. Specific Aim #1 applies the powerful yeast-two-hybrid approach to define and characterize minimal regions of subunit association. Specific Aim #2 answers the question of whether glycosylation influences subunit association in AMPA receptors. Analytical ultracentrifugation experiments in Specific Aim #3 are designed to determine the molecular weight, stoichiometry, and association properties of amino-terminal fragments, which have been shown to self-associated in co- inmmunoprecipitation experiments. Finally, Specific Aim #4 is designed to determine whether AMPA receptor assembly occurs in the endoplasmic reticulum or in another intracellular compartment. In combination with previous data, these experiments will provide a comprehensive description of AMPA receptor biogenesis.