Mutations of the conserved DRS motif in the second intracellular loop of the gonadotropin-releasing hormone receptor affect expression, activation, and internalization
Mutations of the conserved DRS motif in the second intracellular loop of the gonadotropin-releasing hormone receptor affect expression, activation, and internalization
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DOI：
10.1210/me.11.9.1203
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发表时间：
1997-08-01
影响因子：
--
通讯作者：
Catt, KJ
中科院分区：
文献类型：
--
作者：
Arora, KK;Cheng, ZY;Catt, KJ
GnRH受体是G蛋白偶联受体（GPCR）超家族的一个不寻常的成员，具有几个独特的功能。其中之一是保守的DRY基序的变体，其位于大多数GPCR的第三跨膜结构域和第二胞内（2 i）环的连接处。在GnRH受体中，保守三联体的Tyr残基被Ser取代，得到DRS序列。三联体的天冬氨酸和精氨酸残基在几乎所有的GPCR中是高度保守的。在COS-7细胞中表达的野生型和突变型GnRH受体中评价了这些残基的功能重要性。突变体中，天冬氨酸（138）被替换为天冬酰胺或谷氨酸表达差，但表现出显着增加的内化，并表现出增强的肌醇磷酸产生最大的激动剂刺激相比，野生型受体。与此相反，Arg（139）Gln突变体中Arg（139）Gln被Gln、Ala或Ser取代的受体表现出降低的内化，并且Arg（139）Gln突变体的GnRH诱导的磷酸肌醇反应与其低表达水平成比例地显著受损。用Ala取代Ser（140）既不影响内化也不影响信号转导。在另外两种突变体（Ser（151）Ala、Ser（153）Ala和Ser（151）Ala、Ser（153）Ala、Lys（154）Gln、Glu（156）Gln）中评价了2 i环C末端极性氨基酸的作用。这两种突变体都表现出类似于野生型受体的激动剂诱导的磷酸肌醇反应，但表现出增加的受体内化。这种突变分析表明，保守的DRY/S三联体中的天冬氨酸和精氨酸残基作出重要贡献的受体的结构完整性和影响受体的表达，激动剂诱导的激活，和内化。
The GnRH receptor is an unusual member of the G protein-coupled receptor (GPCR) superfamily with several unique features. One of these is a variant of the conserved DRY motif that is located at the junction of the third transmembrane domain and the second intracellular (2i) loop of most GPCRs. In the GnRH receptor, the Tyr residue of the conserved triplet is replaced by Ser, giving a DRS sequence. The aspartate and arginine residues of the triplet are highly conserved in almost all GPCRs. The functional importance of these residues was evaluated in wild type and mutant GnRH receptors expressed in COS-7 cells. Mutants in which Asp(138) was replaced by Asn or Glu were poorly expressed, but showed significantly increased internalization and exhibited augmented inositol phosphate generation to maximal agonist stimulation compared with the wild type receptor. In contrast, receptors in which Arg(139) was substituted With Gin, Ala, or Ser showed reduced internalization, and the GnRH-induced inositol phosphate response for the Arg(139)Gln mutant was significantly impaired in proportion to its low expression lever. Replacing Ser(140) with Ala affected neither internalization nor signal transduction. The role of the polar amino acids at the C terminus of the 2i loop was evaluated in two additional mutants (Ser(151)Ala, Ser(153)Ala, and Ser(151)Ala, Ser(153)Ala, Lys(154)Gln, Glu(156)Gln). Both of these mutants exhibited agonist-induced inositol phosphate responses similar to that of the wild type receptor, but showed increased receptor internalization. This mutational analysis indicates that the conserved Asp and Arg residues in the DRY/S triplet make important contributions to the structural integrity of the receptor and influence receptor expression, agonist-induced activation, and internalization.