GMPPB突变导致抗肌萎缩相关糖蛋白病的机制研究及其在疾病预测与治疗中的应用

GDP-mannose pyrophosphorylase B (GMPPB) catalyzes the formation of GDP-mannose, which is required for the glycosylation of proteins. GMPPB mutations can result in dystroglycanopathy with hypoglycosylated alpha-dystroglycan. However, the underlying mechanism by which GMPPB mutations contribute to the pathogenesis of dystroglycanopathy remains elusive. Furthermore, no effective medical treatments for dystroglycanopathy are currently available. We discovered novel GMPPB mutations in an affected Chinese kid, and the mutant with compromised enzymatic activity fails to rescue the phenotypes in zebrafish disease model, and we also found that the enzymatic activity of GMPPB mutants correlates inversely with the severity of the disease in all known cases, which indicates the involvement of compromised enzymatic activity in the pathogenesis of this disease. In the proposed program, we will further determine the mechanism of compromised enzymatic activity of GMPPB mutants in the pathogenesis of dystroglycanopathy and the severity of the disease. Given the essential role of GDP-mannose catalyzed by GMPPB in glycosylation of alpha-dystroglycan, GMPPB pathway might be an attractive target in the design of therapeutic targets. We will search potential therapeutic approaches targeting GMPPB pathway using zebrafish model. The expected outcomes will not only elucidate the molecular mechanism of GMPPB mutations underlying dystroglycanopathy, but also provide new insights into the prediction of disease due to GMPPB mutations, and into the development of new therapy strategies.

GDP-甘露糖焦磷酸化酶B（GMPPB）催化产生的GDP-甘露糖参与蛋白质糖基化修饰，该基因突变可导致α-肌营养不良蛋白聚糖的糖基化缺陷而引发抗肌萎缩相关糖蛋白病。解析GMPPB突变的致病机制及开发有效的临床筛查与治疗手段是亟待解决的科学问题。前期研究中，我们在抗肌萎缩相关糖蛋白病患者中发现了新的GMPPB突变，其中一个突变可显著降低其酶活性，且该突变体不能恢复斑马鱼疾病模型的表型。我们还发现GMPPB突变体酶活性与患者临床症状的严重程度呈负相关。这些结果表明GMPPB酶活性下降是致病的关键因素。本项目将进一步探索GMPPB突变的致病机制，阐明酶活性下降在疾病发生及决定疾病严重程度中的关键作用。在此基础之上，我们拟以GMPPB催化产生GDP-甘露糖的反应途径为治疗靶标，在斑马鱼疾病模型中探寻潜在的治疗方案。预期结果不仅可以阐释GMPPB突变的致病机理，也将对相关疾病的预测及治疗提供新思路。

先天性糖基化障碍（CDG）是一类蛋白质或脂类糖基化缺乏的代谢性疾病。GDP-甘露糖焦磷酸化酶B（GMPPB），以甘露糖-1-磷酸和GTP为底物，催化GDP-甘露糖的形成，其突变可导致CDG（GMPPB-CDG）。GMPPB-CDG患者临床表现具有异质性，既包括症状较为缓和的肢带型肌营养不良症，先天性肌无力综合症，也包括严重的先天性肌营养不良症。目前已在患者中发现了GMPPB基因的50多个突变，但基因型-表型的相关性仍不明确，GMPPB突变导致疾病发生的机制也尚不清楚，这限制了对此类疾病机制的理解和潜在治疗方案的开发。我们报道了一名先天性糖基化障碍患者。患者表现先天性肌营养不良以及小脑发育异常。利用外显子组测序技术，我们发现患者携带GMPPB基因的复合杂合子突变（c.332T>G (p.Val111Gly)和c.640G>A (p.Gly214Ser))。我们开发了一种测定GMPPB酶活性的方法，发现V111G突变显著降低了GMPPB的酶活性。通过进一步检测17个在患者中鉴定的GMPPB突变体的酶活性，我们发现所有突变体的酶活性均显著下降。为了确定GMPPB在神经肌肉发育过程中的功能，我们以斑马鱼作为模式生物，发现GMPPB突变体的酶活性与斑马鱼的肌肉和神经元表型相关。我们的研究揭示了GMPPB-CDG的发生与GMPPB酶活性下降相关，为该类疾病在临床上的治疗提供了重要参考依据，或可为该类疾病的筛查提供新指标。