A Missense Glial Cells Missing Homolog B (GCMB) Mutation, Asn502His, Causes Autosomal Dominant Hypoparathyroidism
A Missense Glial Cells Missing Homolog B (GCMB) Mutation, Asn502His, Causes Autosomal Dominant Hypoparathyroidism
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DOI：
10.1210/jc.2009-2532
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发表时间：
2010-07-01
影响因子：
5.8
通讯作者：
Thakker, Rajesh V.
中科院分区：
文献类型：
--
作者：
Mirczuk, Samantha M.;Bowl, Michael R.;Thakker, Rajesh V.
内容：神经胶质细胞缺失B（GCMB）是果蝇GCM基因的哺乳动物同源物，编码一个506个氨基酸的甲状旁腺特异性转录因子。迄今为止，只有两个不同的杂合子GCMB突变已被报道在三个kinematis与常染色体显性遗传性甲状旁腺功能减退症。目的：我们的目的是调查一个家庭的常染色体显性遗传性甲状旁腺功能减退症的PTH，CaSR，和GCMB突变。使用荧光显微镜、EMSA和荧光素酶报告基因测定进行功能分析。知情同意书获得了国家伦理委员会批准的协议。结果：DNA序列分析显示，在GCMB的密码子502，改变了野生型天冬酰胺（Asn）组氨酸（His）的A到C颠换。使用GCMB野生型和突变体（Asn 502 His）标记的构建体瞬时转染COS 7细胞的功能研究表明，野生型和突变体蛋白定位于细胞核，并保留了结合GCM-共有DNA识别基序的能力。然而，荧光素酶报告基因分析表明，Asn 502 His突变导致基因反式激活减少。此外，野生型与突变体的共转染没有导致荧光素酶活性的增加，从而证明Asn 502 His突变体的显性负效应，这与常染色体显性遗传一致。我们的研究结果，已经确定了第一个显性错义GCMB突变，有助于增加我们对基因反式激活机制的理解，这是甲状旁腺特异性转录因子功能的先决条件。（临床内分泌代谢杂志95：3512-3516，2010）
Context: Glial cells missing B (GCMB), the mammalian homolog of the Drosophila GCM gene, encodes a 506-amino-acid parathyroid-specific transcription factor. To date, only two different heterozygous GCMB mutations have been reported in three kindreds with autosomal dominant hypoparathyroidism.Objective: Our objective was to investigate a family with autosomal dominant hypoparathyroidism for PTH, CaSR, and GCMB mutations.Methods: Leukocyte DNA was used with exon-specific primers for PCR amplification and the DNA sequences of the PCR products determined. Functional analyses using fluorescence microscopy, EMSAs, and luciferase reporter assays were undertaken. Informed consent was obtained using protocols approved by a national ethical committee.Results: DNA sequence analysis revealed an A to C transversion at codon 502 of GCMB, which altered the wild-type asparagine (Asn) to histidine (His). Functional studies, using transient transfections of COS7 cells with GCMB wild-type and mutant (Asn502His) tagged constructs, demonstrated that the wild-type and mutant proteins localized to the nucleus and retained the ability to bind the GCM-consensus DNA recognition motif. However, a luciferase reporter assay demonstrated that the Asn502His mutation resulted in a reduction in gene transactivation. Moreover, cotransfection of the wild-type with mutant did not lead to an increase in luciferase activity, thereby demonstrating a dominant-negative effect of the Asn502His mutant that would be consistent with an autosomal dominant inheritance.Conclusion: Our results, which have identified the first dominant missense GCMB mutation, help to increase our understanding of the mechanism underlying gene transactivation that is a prerequisite for the function of this parathyroid gland-specific transcription factor. (J Clin Endocrinol Metab 95: 3512-3516, 2010)