SPLICE VARIANTS OF THE N-METHYL-D-ASPARTATE RECEPTOR NR1 IDENTIFY DOMAINS INVOLVED IN REGULATION BY POLYAMINES AND PROTEIN-KINASE-C
SPLICE VARIANTS OF THE N-METHYL-D-ASPARTATE RECEPTOR NR1 IDENTIFY DOMAINS INVOLVED IN REGULATION BY POLYAMINES AND PROTEIN-KINASE-C
复制标题
DOI：
10.1073/pnas.90.14.6731
复制
发表时间：
1993-07-15
影响因子：
11.1
通讯作者：
ZUKIN, RS
中科院分区：
文献类型：
--
作者：
DURAND, GM;BENNETT, MVL;ZUKIN, RS
N-甲基-D-天冬氨酸 (NMDA) 受体 NR1 基因编码 RNA，该 RNA 可以选择性剪接以产生至少七个变体。这些变异是由三个外显子的剪接引起的。一种编码 N 端结构域中的 21 个氨基酸插入片段，两种编码 C 端结构域中的 37 和 38 个氨基酸的相邻序列。第二个 C 末端外显子的剪接会删除一个终止密码子，并在到达第二个终止密码子之前产生一个额外的开放阅读框，编码 22 个氨基酸的不相关序列。我们通过存在或不存在三个选择性剪接的外显子（从 5' 到 3'）来表示 NR1 变体；因此，NR1(111) 具有全部三个外显子，NR1(000) 没有，而 NR1(100) 仅具有 N 末端外显子。我们在这里报告了非洲爪蟾卵母细胞中表达的 NR1 受体的六种剪接变体的电生理学特征。缺乏N末端外显子的NR1受体（NR1（000）、NR1（010）和NR1（011））表现出对NMDA相对较高的亲和力（EC50几乎等于13μM）并且对精胺具有显着的增强作用。相反，那些带有 N 末端插入片段的受体变体（NR1(100)、NR1(101) 和 NR1(111)）显示出较低的激动剂亲和力，并且在饱和甘氨酸时几乎没有或没有精胺增强作用。所有六种变体均在低甘氨酸下显示出精胺增强作用，并在更负电位下显示出精胺抑制作用。仅 C 端结构域不同的变体在激动剂亲和力和精胺增强方面几乎没有差异。这些发现表明，N 端插入物要么参与激动剂和多胺结合域，要么间接修饰它们的构象。剪接变体的不同之处在于它们被蛋白激酶 C (PKC) 激活剂增强的程度从 3 到 20 倍。 N 端插入片段的存在和 C 端序列的缺失增加了 PKC 的增强作用。这些发现确定了单独的多肽结构域对多胺和 PKC 调节的贡献，并为亚基组成决定 NMDA 受体功能特性的概念提供了进一步的支持。
The N-methyl-D-aspartate (NMDA) receptor NR1 gene encodes RNA that is alternatively spliced to generate at least seven variants. The variants arise from splicing in or out of three exons; one encodes a 21-amino acid insert in the N-terminal domain, and two encode adjacent sequences of 37 and 38 amino acids in the C-terminal domain. Splicing out of the second C-terminal exon deletes a stop codon and results in an additional open reading frame encoding an unrelated sequence of 22 amino acids before arriving at a second stop codon. We denote the NR1 variants by the presence or absence of the three alternatively spliced exons (from 5' to 3'); thus, NR1(111) has all three exons, NR1(000) has none, and NR1(100) has only the N-terminal exon. We report here electrophysiological characterization of six splice variants of the NR1 receptor expressed in Xenopus oocytes. NR1 receptors that lacked the N-terminal exon (NR1(000), NR1(010), and NR1(011)) exhibited a relatively high affinity for NMDA (EC50 almost-equal-to 13 muM) and marked potentiation by spermine. In contrast, those receptor variants with the N-terminal insert (NR1(100), NR1(101), and NR1(111)) showed a lower agonist affinity and little or no spermine potentiation at saturating glycine. All six variants showed spermine potentiation at low glycine and inhibition by spermine at more negative potentials. Variants differing only in the C-terminal domain differed little in agonist affinity and spermine potentiation. These findings indicate that the N-terminal insert either participates in agonist and polyamine binding domains or indirectly modifies their conformations. The splice variants differed in the extent to which they could be potentiated by activators of protein kinase C (PKC) from 3- to 20-fold. Presence of the N-terminal insert and absence of the C-terminal sequences increased potentiation by PKC. These findings identify the contributions of the separate polypeptide domains to modulation by polyamines and PKC and provide further support for the concept that subunit composition determines functional properties of NMDA receptors.