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非洲爪蟾胚胎间隙连接蛋白的克隆与表达

Cloning and expression of a Xenopus embryonic gap junction protein.

作者信息

Ebihara L, Beyer E C, Swenson K I, Paul D L, Goodenough D A

机构信息

Department of Pharmacology, Columbia University, New York, NY 10032.

出版信息

Science. 1989 Mar 3;243(4895):1194-5. doi: 10.1126/science.2466337.

Abstract

Gap junctions in the early amphibian embryo may play a fundamental role in the regulation of differentiation by mediating the cell-to-cell transfer of chemical signals. A complementary DNA encoding a gap junction present in Xenopus oocytes and early embryos has now been cloned and sequenced. This protein sequence is homologous to the well-characterized gap junction structural proteins rat connexin32 and connexin43. RNA blot analysis of total Xenopus oocyte RNA showed hybridization to a single 1.6-kilobase band. This messenger RNA is abundant in oocytes, decreases to levels below the sensitivity of our assay by stage 15 (18 hours), and is not detectable in RNA from a number of adult organs. To confirm that the oocyte cDNA encodes a gap junction channel, the protein was over expressed in Xenopus oocytes by injection of RNA synthesized in vitro. Pairs of RNA-injected oocytes formed many more time- and voltage-sensitive cell-cell channels than water-injected pairs.

摘要

早期两栖类胚胎中的间隙连接可能通过介导化学信号的细胞间传递,在分化调控中发挥重要作用。现在已经克隆并测序了一种编码非洲爪蟾卵母细胞和早期胚胎中存在的间隙连接的互补DNA。该蛋白质序列与特征明确的间隙连接结构蛋白大鼠连接蛋白32和连接蛋白43同源。对非洲爪蟾卵母细胞总RNA的RNA印迹分析显示与一条单一的1.6千碱基条带杂交。这种信使RNA在卵母细胞中含量丰富,到第15阶段(18小时)时降至低于我们检测灵敏度的水平,并且在许多成体器官的RNA中无法检测到。为了证实卵母细胞cDNA编码间隙连接通道,通过注射体外合成的RNA在非洲爪蟾卵母细胞中过量表达该蛋白质。注射RNA的卵母细胞对形成的时间和电压敏感的细胞间通道比注射水的卵母细胞对多得多。

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