Manipal Academy of Higher Education (MAHE), Tiger Circle Road, Madhav Nagar, Manipal 576 104, Karnataka, India.
Department of Laboratory Medicine and Pathology, Mayo Clinic, 200 First Street SW, Rochester, Minnesota 55905, United States.
Glycobiology. 2024 Sep 30;34(11). doi: 10.1093/glycob/cwae076.
SRD5A3-CDG is a congenital disorder of glycosylation (CDG) resulting from pathogenic variants in SRD5A3 and follows an autosomal recessive inheritance pattern. The enzyme encoded by SRD5A3, polyprenal reductase, plays a crucial role in synthesizing lipid precursors essential for N-linked glycosylation. Despite insights from functional studies into its enzymatic function, there remains a gap in understanding global changes in patient cells. We sought to identify N-glycoproteomic and proteomic signatures specific to SRD5A3-CDG, potentially aiding in biomarker discovery and advancing our understanding of disease mechanisms. Using tandem mass tag (TMT)-based relative quantitation, we analyzed fibroblasts derived from five patients along with control fibroblasts. N-glycoproteomics analysis by liquid chromatography-tandem mass spectrometry (LC-MS/MS) identified 3,047 glycopeptides with 544 unique N-glycosylation sites from 276 glycoproteins. Of these, 418 glycopeptides showed statistically significant changes with 379 glycopeptides decreased (P < 0.05) in SRD5A3-CDG patient-derived samples. These included high mannose, complex and hybrid glycan-bearing glycopeptides. High mannose glycopeptides from protocadherin Fat 4 and integrin alpha-11 and complex glycopeptides from CD55 were among the most significantly decreased glycopeptides. Proteomics analysis led to the identification of 5,933 proteins, of which 873 proteins showed statistically significant changes. Decreased proteins included cell surface glycoproteins, various mitochondrial protein populations and proteins involved in the N-glycosylation pathway. Lysosomal proteins such as N-acetylglucosamine-6-sulfatase and procathepsin-L also showed reduced levels of phosphorylated mannose-containing glycopeptides. Our findings point to disruptions in glycosylation pathways as well as energy metabolism and lysosomal functions in SRD5A3-CDG, providing clues to improved understanding and management of patients with this disorder.
SRD5A3-CDG 是一种先天性糖基化障碍（CDG），由 SRD5A3 中的致病性变异引起，呈常染色体隐性遗传模式。SRD5A3 编码的多烯醇还原酶对于合成 N-连接糖基化所需的脂质前体至关重要。尽管对其酶功能的功能研究提供了一些见解，但对患者细胞中全局变化的理解仍存在差距。我们试图确定 SRD5A3-CDG 特有的 N-糖蛋白组学和蛋白质组学特征，这可能有助于发现生物标志物并深入了解疾病机制。我们使用串联质量标签（TMT）相对定量法分析了来自五名患者和对照患者的成纤维细胞。通过液相色谱-串联质谱（LC-MS/MS）的 N-糖蛋白组学分析，从 276 种糖蛋白中鉴定出 3047 种糖肽，其中包含 544 个独特的 N-糖基化位点。其中，418 种糖肽的变化具有统计学意义，379 种糖肽（SRD5A3-CDG 患者样本中降低，P < 0.05）。这些糖肽包括高甘露糖、复合和杂合糖基化的糖肽。原钙粘蛋白 Fat 4 和整合素 alpha-11 的高甘露糖糖肽以及 CD55 的复合糖肽是降低最显著的糖肽之一。蛋白质组学分析鉴定出 5933 种蛋白质，其中 873 种蛋白质发生了统计学上的显著变化。降低的蛋白质包括细胞表面糖蛋白、各种线粒体蛋白群以及参与 N-糖基化途径的蛋白质。溶酶体蛋白，如 N-乙酰氨基葡萄糖-6-硫酸酯酶和前组织蛋白酶 L，也显示出磷酸化含甘露糖的糖肽水平降低。我们的研究结果表明，SRD5A3-CDG 中存在糖基化途径以及能量代谢和溶酶体功能的中断，为改善对这种疾病患者的理解和管理提供了线索。
