Cloning of a thyroid hormone-responsive Rana catesbeiana c-erbA-beta gene.
Cloning of a thyroid hormone-responsive Rana catesbeiana c-erbA-beta gene.
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甲状腺激素响应性林蛙 c-erbA-beta 基因的克隆。
DOI：
10.1002/dvg.1020150405
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发表时间：
1994
期刊：
影响因子：
--
通讯作者：
Galton,VA
中科院分区：
文献类型：
--
作者：
Davey,JC;Schneider,MJ;Galton,VA
两种类型的甲状腺激素受体（c - erbA）基因已在哺乳动物和较低等物种（包括鸡和两栖动物）中被鉴定出来。这两个基因位于不同的染色体上，分别被命名为TRα和TRβ。我们之前已经报道了从红毛斑斑蝶（RC15）组织中克隆TRβ cDNA，现在我们报道了从该物种中克隆TRβ cDNA。克隆策略采用聚合酶链反应（PCR），引物基于theX的序列。laevisTRβ cDNA （XenTRβ）和RCTRβ基因组克隆，该克隆与XenTRβ相似，包含一些开放阅读框的3 ‘端以及3 ’‐未翻译序列。在核苷酸和氨基酸水平上，克隆的RCTRβ cDNA与XenTRβ的同源性分别为90%和98%，与RC15的同源性分别为72%和76%。经过体外转录和翻译，该cDNA编码了一个48千尔顿的蛋白，该蛋白以高亲和力（平均Kd: 0.032 nM）结合3,5,3′‐三碘甲状腺原氨酸（T3）。利用258 bp的RCTRβ cDNA片段作为探针，对不同变态阶段蝌蚪和成年蛙的总RNA或多(A)+RNA样本进行了凹槽印迹分析，以确定是否存在trβ特异性转录物。这段cDNA不能与RC15的相应片段杂交。为了证实先前的发现，在蝌蚪红细胞（rbc）的RNA中未检测到β特异性转录物，在成年青蛙的红细胞中也未发现。然而，在13期蝌蚪的尾巴、皮肤和肝脏的RNA中检测到β特异性转录物，并且在这些组织中的水平在变质高潮期间大大增加。在成年蛙的所有研究组织中，该水平都是最低的。在其他使用12期蝌蚪和逆转录聚合酶链反应的研究中，β特异性转录物在尾巴、皮肤、肾脏、腿、心脏、肠和眼睛中很容易检测到，但在肝脏、大脑和红细胞中很少或不存在。注射T3后2天内，所有组织中β特异性转录物的水平均显著升高。这些发现有力地表明，RCTRβ基因的表达在RC蝌蚪的大多数组织中受到发育和/或甲状腺激素的调节。©1994 Wiley‐Liss, Inc。
Two types of thyroid hormone receptor (c‐erbA) gene have been identified in mammals and in lower species including chickens and the amphibianXenopus laevis.The two genes are located on different chromosomes and have been named TRα and TRβ We have described previously the cloning of a TRβ cDNA fromRana catesbeiana(RC) tissues (RC15) and we now report the cloning of a TRβ cDNA from this species. The cloning strategy employed utilized the polymerase chain reaction (PCR), with primers based on the sequences of theX. laevisTRβ cDNA (XenTRβ) and an RCTRβ genomic clone, which, by analogy with XenTRβ, contains some of the 3′ end of the open reading frame together with 3′‐untranslated sequences. At the nucleotide and amino acid levels, respectively, the cloned RCTRβ cDNA is 90% and 98% homologous with XenTRβ, and 72% and 76% homologous with RC15. Following in vitro transcription and translation, the cDNA was shown to encode a 48 kilodalton protein which binds 3,5, 3′‐triiodothyronine (T3) with high affinity (mean Kd: 0.032 nM). Samples of total or poly(A)+RNA from tadpoles at different stages of metamorphosis and from adult frogs were analyzed for the presence ofTRβ‐specific transcripts by slot blot analysis using as probe a 258 bp section of the RCTRβ cDNA. This section of the cDNA does not hybridize to the corresponding section of RC15. In confirmation of previous findings, β‐specific transcripts were not detected in RNA from tadpole red blood cells (RBCs) and none was found in RBCs from adult frogs. However, β‐specific transcripts were detected in RNA from tail, skin, and liver of stage XIII tadpoles, and the levels were greatly increased in these tissues during metamorphic climax. In the adult frog, the level was minimal in all tissues studied. In other studies using stage XII tadpoles and a reverse transcription‐polymerase chain reaction assay, β‐specific transcripts were readily detected in tail, skin, kidney, leg, heart, intestine, and eye but minimal or absent in liver, brain, and RBCs. Within 2 days of injection of T3, the level of β‐specific transcripts was markedly increased in all tissues. These findings strongly suggest that the expression of the RCTRβ gene is regulated developmentally and/or by thyroid hormone in most tissues of the RC tadpole. © 1994 Wiley‐Liss, Inc.
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