A Functional P2X7 Splice Variant with an Alternative Transmembrane Domain 1 Escapes Gene Inactivation in P2X7 Knock-out Mice
A Functional P2X7 Splice Variant with an Alternative Transmembrane Domain 1 Escapes Gene Inactivation in P2X7 Knock-out Mice
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DOI：
10.1074/jbc.m109.033134
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发表时间：
2009-09-18
影响因子：
4.8
通讯作者：
Soto, Florentina
中科院分区：
文献类型：
--
作者：
Nicke, Annette;Kuan, Yung-Hui;Soto, Florentina
ATP激活的P2 X7受体通道参与免疫功能和炎性疼痛，是重要的药物靶点。在这里，我们描述了一个新的P2 X7剪接变异体（P2 X7（k）），含有一个替代的细胞内N端和第一个跨膜结构域编码的一个新的外显子1在啮齿动物P2 rx 7基因。在HEK 293细胞中表达的大鼠同种型的全细胞膜片钳记录显示，与P2 X7（a）受体相比，对激动剂Bz-ATP的敏感性高8倍，失活动力学慢得多。非洲爪蟾卵母细胞的渗透性测量显示，激活后立即对N-甲基-D-葡糖胺具有高渗透性，这表明P2 X7（k）通道在打开后组成性扩张。激动剂诱导的染料摄取率和HEK细胞膜起泡率也增加。PCR分析和通过SDS-PAGE和BN-PAGE的生化分析表明，P2 X7（k）变体在一个可用的P2 X7（-/-）小鼠品系中逃避基因缺失，并且在脾脏中强烈表达。总之，我们描述了一种新的P2 X7亚型，具有独特的功能特性，有助于P2 X7受体信号的多样性。它在P2 X7（-/-）菌株中的存在对于我们理解这种受体在健康和疾病中的作用具有重要意义。
The ATP-activated P2X7 receptor channel is involved in immune function and inflammatory pain and represents an important drug target. Here we describe a new P2X7 splice variant (P2X7(k)), containing an alternative intracellular N terminus and first transmembrane domain encoded by a novel exon 1 in the rodent P2rx7 gene. Whole cell patch clamp recordings of the rat isoform expressed in HEK293 cells revealed an 8-fold higher sensitivity to the agonist Bz-ATP and much slower deactivation kinetics when compared with the P2X7(a) receptor. Permeability measurements in Xenopus oocytes show a high permeability for N-methyl-D-glucamine immediately upon activation, suggesting that the P2X7(k) channel is constitutively dilated upon opening. The rates of agonist-induced dye uptake and membrane blebbing in HEK cells were also increased. PCR analyses and biochemical analysis by SDS-PAGE and BN-PAGE indicate that the P2X7(k) variant escapes gene deletion in one of the available P2X7(-/-) mice strains and is strongly expressed in the spleen. Taken together, we describe a novel P2X7 isoform with distinct functional properties that contributes to the diversity of P2X7 receptor signaling. Its presence in one of the P2X7(-/-) strains has important implications for our understanding of the role of this receptor in health and disease.