Using sno-lncRNAs as potential markers for Prader-Willi syndrome diagnosis

The genetic disorder Prader-Willi syndrome (PWS) is mainly caused by the loss of multiple paternally expressed genes in chromosome 15q11-q13 (the PWS region). Early diagnosis of PWS is essential for timely treatment, leading to effectively easing some clinical symptoms. Molecular approaches for PWS diagnosis at the DNA level are available, but the diagnosis of PWS at the RNA level has been limited. Here, we show that a cluster of paternally transcribed snoRNA-ended long noncoding RNAs (sno-lncRNAs, sno-lncRNA1–5) derived from the SNORD116 locus in the PWS region can serve as diagnostic markers. In particular, quantification analysis has revealed that 6,000 copies of sno-lncRNA3 are present in 1 μL whole blood samples from non-PWS individuals. sno-lncRNA3 is absent in all examined whole blood samples of 8 PWS individuals compared to 42 non-PWS individuals and dried blood samples of 35 PWS individuals compared to 24 non-PWS individuals. Further developing a new CRISPR-MhdCas13c system for RNA detection with a sensitivity of 10 molecules per μL has ensured sno-lncRNA3 detection in non-PWS, but not PWS individuals. Together, we suggest that the absence of sno-lncRNA3 represents a potential marker for PWS diagnosis that can be detected by both RT-qPCR and CRISPR-MhdCas13c systems with only microlitre amount of blood samples. Such an RNA-based sensitive and convenient approach may facilitate the early detection of PWS.

帕拉德尔-威利综合症（Prader-Willi syndrome，简称PWS）主要由于15q11-q13染色体（PWS区域）上多个父系表达基因的缺失所致。PWS的早期诊断对于及时治疗至关重要，有助于有效缓解部分临床症状。目前，PWS的DNA水平分子诊断方法已得到应用，但在RNA水平的诊断则受到了限制。在本研究中，我们发现PWS区域SNORD116位点起源的一组父系转录的snoRNA末端长非编码RNA（sno-lncRNAs，包括sno-lncRNA1至sno-lncRNA5）可以作为诊断标志物。特别是，定量分析显示，非PWS个体的1 μL全血样本中存在6,000个sno-lncRNA3拷贝。与42名非PWS个体和35名PWS个体的干血样本相比，8名PWS个体的所有全血样本中均未检测到sno-lncRNA3。进一步开发了一种具有每微升10个分子灵敏度的CRISPR-MhdCas13c系统用于RNA检测，确保了非PWS个体中sno-lncRNA3的检测，但在PWS个体中并未检测到。综上所述，sno-lncRNA3的缺失可能成为PWS诊断的潜在标志物，且可通过RT-qPCR和CRISPR-MhdCas13c系统在仅需微量血液样本的情况下进行检测。这种基于RNA的敏感且便捷的方法可能有助于PWS的早期发现。