An Expanded Spectrum for Congenital Disorders of Glycosylation

描述（由申请人提供）：该项目将基础科学与患者相关的转化研究相结合，因为它确定了损害蛋白质n -糖基化的罕见遗传疾病的原因。今天，已知有39个基因引起先天性糖基化疾病（CDG），但我们知道许多导致CDG的基因仍未被发现。该建议使用一种新的、公正的、前瞻性的、功能性的方法来识别这些新基因。具体目标在基础科学和患者的遗传缺陷之间建立了一座双向的桥梁。它将识别新的基因，以指导和告知医生在低于1000美元的基因组时代对未来CDG患者的诊断。CDGs影响每个器官系统，具有一系列令人惊叹的不同临床表现，跨越每个医学专业。临床表现不能确定该基因，然而，几乎所有当前CDG患者都有异常的血清转铁蛋白（Tf）糖基化生物标志物。目前还没有类似tf的细胞生物标志物可用于评估疑似糖基化缺陷患者的糖基化或补体候选基因。为了填补这一空白，我们构建了一个含有n -糖基化位点（Glyc-ER-GFP）的er -保留GFP，只有当该位点未被占用时才允许荧光。这为糖基化受损的细胞提供了高度敏感的n-聚糖生物传感器：它们会发光。这种新的细胞生物标志物可以通过筛选一个完善的人类全基因组siRNA敲低文库来识别新的n -糖基化基因。已知的CDG基因作为筛查的阳性对照。二级糖基化生物标志物和生化分析可以证实新基因参与n -糖基化。这种方法有即时和长期的回报。功能糖基化试验可以识别基因，以查询今天的CDG患者的未知缺陷。未来，在全外显子组（基因组）测序管道中显示这些基因突变的受试者将知道它影响n -糖基化途径。新发现的基因也将扩大我们目前对蛋白质糖基化的理解。

DESCRIPTION (provided by applicant): This project merges basic science with patient-relevant translational research because it identifies the causes of rare genetic diseases that impair protein N-glycosylation. Today, 39 genes are known to cause Congenital Disorders of Glycosylation (CDG), but we know that many CDG-causing genes remain undiscovered. This proposal uses a new, unbiased, forward-reaching, functional approach to identify these new genes. The Specific Aims build a bi-directional bridge between fundamental science and genetic defects in patients today. It will identify new genes to guide and inform physicians in their diagnosis of tomorrow's CDG patients in the sub-$1000-genome era. CDGs affect every organ system with a breath-taking array of variable clinical presentations that cross every medical specialty. Clinical presentations do not identify the gene, however, nearly all current CDG patients have an abnormal serum transferrin (Tf) glycosylation biomarker. There is no Tf-like cellular biomarker that can be used to assess glycosylation or complement candidate genes in suspected glycosylation-deficient patients. To fill this void, we constructed an ER-retained GFP with an N-glycosylation site (Glyc-ER-GFP) that allows fluorescence only when that site is unoccupied. This provides a highly sensitive N-glycan-biosensor for cells with impaired glycosylation: They glow. This novel cellular biomarker can identify new N-glycosylation genes by screening a well-established human genome-wide siRNA knockdown library for cells that glow. The known CDG genes serve as positive controls for the screening. Secondary glycosylation biomarkers and biochemical assays can confirm the novel gene's involvement in N-glycosylation. This approach has immediate and long-term payoffs. The functional glycosylation assay can identify genes to query today's CDG patients with unknown defects. In the future, subjects in the whole exome (genome) sequencing pipeline who show mutations in these genes will know it impacts the N-glycosylation pathway. The newly discovered genes will also expand the dimensions of our current understanding about protein glycosylation.