A G220V substitution within the N-terminal transcription regulating domain of HOXD13 causes a variant synpolydactyly phenotype
A G220V substitution within the N-terminal transcription regulating domain of HOXD13 causes a variant synpolydactyly phenotype
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DOI：
10.1093/hmg/ddn410
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发表时间：
2009-03-01
影响因子：
3.5
通讯作者：
Zappavigna, Vincenzo
中科院分区：
文献类型：
--
作者：
Fantini, Sebastian;Vaccari, Giulia;Zappavigna, Vincenzo
HoxD基因簇的5'成员（同源群9-13）对正确的脊椎动物肢体模式至关重要。已发现HOXD13基因突变可导致人类多指畸形（SPD）和其他肢体畸形。我们报告了在一个希腊家族中鉴定出一种由HOXD13的220位（G220V）的甘氨酸取代缬氨酸的新型错义突变引起的SPD变体形式。这种突变代表了位于HOXD13同源结构域外的氨基酸的第一次替换，导致自节肢体畸形。我们在分子水平上对这一突变进行了表征，发现G220V取代导致HOXD13结合DNA和调节转录的能力显著受损。发现HOXD13（G220V）缺乏通过HOXD13响应的调控元件激活和抑制转录。根据这些结果，利用逆转录病毒介导的小鸡肢体错误表达，对HOXD13和HOXD13（G220V）在体内的活性进行比较，发现G220V突变削弱了HOXD13干扰近端肢体骨骼元件发育和异位激活Hand2靶基因转录的能力。此外，我们还发现G220V突变损害了HOXD13蛋白在细胞内的稳定性，并导致其以微妙聚集体的形式部分积聚在细胞质中。综上所述，我们的研究结果表明，G220V替代不会产生显性负作用或功能获得，而是代表了显性功能缺失突变，揭示了人类HOXD13的单倍不足。
The 5' members of the HoxD gene cluster (paralogous groups 9-13) are crucial for correct vertebrate limb patterning. Mutations in the HOXD13 gene have been found to cause synpolydactyly (SPD) and other limb malformations in human. We report the identification in a Greek family of a variant form of SPD caused by a novel missense mutation that substitutes glycine for valine in position 220 (G220V) of HOXD13. This mutation represents the first substitution of an amino acid located outside of the HOXD13 homeodomain that causes autopodal limb malformations. We have characterized this mutation at the molecular level and found that the G220V substitution causes a significant impairment of the capacity of HOXD13 to bind DNA and regulate transcription. HOXD13(G220V) was found to be deficient in both activating and repressing transcription through HOXD13-responsive regulatory elements. In accordance with these results, a comparison of the activities of HOXD13 and HOXD13(G220V) in vivo, using retrovirus-mediated misexpression in developing chick limbs, showed that the G220V mutation impairs the capacity of HOXD13 to perturb the development of proximal limb skeletal elements and to ectopically activate the transcription of the Hand2 target gene. We moreover show that the G220V mutation compromises the stability of the HOXD13 protein within cells and causes its partial accumulation in the cytosol in the form of subtle aggregates. Taken together, our results establish that the G220V substitution does not produce a dominant-negative effect or a gain-of-function, but represents a dominant loss-of-function mutation revealing haploinsufficiency of HOXD13 in human.