GlyT1 encephalopathy: Characterization of presumably disease causing GlyT1 mutations
GlyT1 encephalopathy: Characterization of presumably disease causing GlyT1 mutations
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DOI：
10.1016/j.neuint.2020.104813
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发表时间：
2020-10-01
影响因子：
4.2
通讯作者：
Eulenburg, V
中科院分区：
文献类型：
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作者：
Hauf, K.;Barsch, L.;Eulenburg, V
甘氨酸是中枢神经系统尾侧区域的主要抑制性神经递质。细胞外甘氨酸浓度由两种高亲和力、大容量转运蛋白GlyT 1和GlyT 2协同调节。这两种蛋白质分别由单个基因SLC 6A 9和SLC 6A 5编码。编码GlyT 2的SLC 6A 5基因内的突变已被证明是人类复杂神经肌肉疾病-神经丛过度（OMIM #614618）的病因。相比之下，编码GlyT 1的SLC 6A 9基因内的突变与GlyT 1脑病（OMIM #601019）相关，这是一种引起严重的出生后呼吸缺陷、肌肉张力减退和关节弯曲的疾病。然而，尚未分析各自GlyT 1突变对转运蛋白功能的影响。在这项研究中，我们提出了三个先前发表的GlyT 1突变的功能特征，两个突变预测会导致GlyT 1（GlyT 1（Q573*）和Gly（T1 K310 F +fs*31））的截短和一个预测会导致跨膜结构域7的转运蛋白（GlyT 1（S407 G）），这与GlyT 1脑病。此外，表征的一种新的突变，预测引起跨膜结构域1（GlyT 1（V118 M））内的氨基酸交换确定在两个胎儿显示增加颈部的不透明性和arthrogriposis在常规超声扫描证明。我们表明，在重组系统中的两个大概截断突变导致细胞内保留GlyT 1蛋白缺乏细胞内C末端结构域。在这两种情况下，这种截短的蛋白质没有显示出任何残留的运输活性。点突变hGlyT 1（S407 G）和hGlyT 1（V118 M）被正确处理，但显示出严重降低的活性，因此构成了体内功能性敲除。总之，我们的数据表明，所有分析的GlyT 1突变，已确定在GlyT 1脑病患者造成严重损害的转运功能。这与GlyT 1功能丧失确实是疾病表型的原因这一观点一致。
Glycine constitutes a major inhibitory neurotransmitter predominantly in caudal regions of the CNS. The extracellular glycine concentration is regulated synergistically by two high affinity, large capacity transporters GlyT1 and GlyT2. Both proteins are encoded by single genes SLC6A9 and SLC6A5, respectively. Mutations within the SLC6A5 gene encoding for GlyT2 have been demonstrated to be causative for hyperekplexia (OMIM #614618), a complex neuromuscular disease, in humans. In contrast, mutations within the SLC6A9 gene encoding for GlyT1 have been associated with GlyT1 encephalopathy (OMIM #601019), a disease causing severe postnatal respiratory deficiency, muscular hypotonia and arthrogryposis. The consequences of the respective GlyT1 mutations on the function of the transporter protein, however, have not yet been analysed. In this study we present the functional characterisation of three previously published GlyT1 mutations, two mutations predicted to cause truncation of GlyT1 (GlyT1(Q573*) and Gly(T1K310F+fs*31)) and one predicted to cause an amino acid exchange within transmembrane domain 7 of the transporter (GlyT1(S407G)), that are associated with GlyT1 encephalopathy. Additionally, the characterization of a novel mutation predicted to cause an amino acid exchange within transmembrane domain 1 (GlyT1(V118M)) identified in two fetuses showing increased nuchal translucency and arthrogryposis in routine ultrasound scans is demonstrated. We show that in recombinant systems the two presumably truncating mutations resulted in an intracellular retained GlyT1 protein lacking the intracellular C terminal domain. In both cases this truncated protein did not show any residual transport activity. The point mutations, hGlyT1(S407G) and hGlyT1(V118M), were processed correctly, but showed severely diminished activity, thus constituting a functional knock-out in-vivo. Taken together our data demonstrate that all analysed mutations of GlyT1 that have been identified in GlyT1 encephalopathy patients cause severe impairment of transporter function. This is consistent with the idea that loss of GlyT1 function is indeed causal for the disease phenotype.