The R271W mutant form of Pit-1 does not act as a dominant inhibitor of Pit-1 action to activate the promoters of GH and prolactin genes.
The R271W mutant form of Pit-1 does not act as a dominant inhibitor of Pit-1 action to activate the promoters of GH and prolactin genes.
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DOI：
10.1530/eje.0.1480619
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发表时间：
2003-06
影响因子：
5.8
通讯作者：
M. Kishimoto;Y. Okimura;M. Fumoto;G. Iguchi;K. Iida;H. Kaji;K. Chihara
中科院分区：
文献类型：
--
作者：
M. Kishimoto;Y. Okimura;M. Fumoto;G. Iguchi;K. Iida;H. Kaji;K. Chihara
目的垂体特异性转录因子Pit-1的遗传异常在一些GH、催乳素（PRL）和TSH缺乏症患者中已被报道。最常见的突变是在Pit-1基因的一个等位基因的密码子271 （R271W）中将精氨酸变为色氨酸。根据之前的体外表达研究，R271W作为野生型的显性阴性抑制剂激活GH启动子。然而，也有报道称，携带这种突变的健康携带者应该受到R271W的显性负作用的影响。本研究的目的是更详细地阐明这种突变形式的Pit-1的功能。方法采用瞬时转染法研究R271W在COS7细胞中对pit -1相关基因表达的转录活性。将1.8 kb大鼠GH、0.6 kb大鼠PRL或1.9 kb大鼠PRL 5'侧区插入荧光素酶报告基因上游，用于R271W的功能分析。另一个包含7个Pit-1应答元件的报告基因也被使用。同样的实验也用JEG3和CHO细胞进行。结果未证实R271W对野生型Pit-1的显性负作用。此外，我们的表达研究表明，R271W可以激活GH和PRL基因的启动子，其水平与野生型相似。结论结合已报道的表型正常病例中该突变的证据，我们的研究结果否定了R271W与联合垂体激素缺乏症之间的关系。
OBJECTIVE Genetic abnormalities of the pituitary specific transcription factor, Pit-1, have been reported in several patients with GH, prolactin (PRL) and TSH deficiencies. The most common is a mutation altering an arginine to a tryptophan in codon 271 (R271W) in one allele of the Pit-1 gene. According to the previous in vitro expression study, R271W acted as a dominant negative inhibitor of the wild type to activate the GH promoter. However, healthy carriers with this mutation, who should be affected by the dominant negative effect of R271W, have also been reported. The aim of this study was to clarify in more detail the function of this mutant form of Pit-1. METHODS Transcriptional activity of R271W for the expression of Pit-1-associated genes was investigated in COS7 cells with the aid of transient transfection assays. The 1.8 kb rat GH, 0.6 kb rat PRL or 1.9 kb rat PRL 5'-flanking regions were inserted upstream of the luciferase reporter gene and were used for functional analysis of R271W. Another reporter gene containing seven Pit-1 responsive elements was also used. The same experiments were also performed using JEG3 and CHO cells. RESULTS We could not confirm the dominant negative effect of R271W on wild type Pit-1. Furthermore, our expression study revealed that R271W could activate the promoters of GH and PRL genes to levels similar to the wild type. CONCLUSION Taken together with the evidence that phenotypically normal cases have been reported with this mutation, our results deny the relationship between R271W and combined pituitary hormone deficiency.