Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada, T6G 2G2.
Biomedical Chemistry Institute, Department of Chemistry, New York University, New York, New York 10003, United States.
ACS Chem Biol. 2021 Oct 15;16(10):1900-1907. doi: 10.1021/acschembio.1c00247. Epub 2021 Jun 4.
MicroRNAs (miRNAs, miRs) finely tune protein expression and target networks of hundreds to thousands of genes that control specific biological processes. They are critical regulators of glycosylation, one of the most diverse and abundant post-translational modifications. In recent work, miRs have been shown to predict the biological functions of glycosylation enzymes, leading to the "miRNA proxy hypothesis" which states, "if a miR drives a specific biological phenotype..., the targets of that miR will drive the same biological phenotype." Testing of this powerful hypothesis is hampered by our lack of knowledge about miR targets. Target prediction suffers from low accuracy and a high false prediction rate. Herein, we develop a high-throughput experimental platform to analyze miR-target interactions, miRFluR. We utilize this system to analyze the interactions of the entire human miRome with beta-3-glucosyltransferase (B3GLCT), a glycosylation enzyme whose loss underpins the congenital disorder Peters' Plus Syndrome. Although this enzyme is predicted by multiple algorithms to be highly targeted by miRs, we identify only 27 miRs that downregulate B3GLCT, a >96% false positive rate for prediction. Functional enrichment analysis of these validated miRs predicts phenotypes associated with Peters' Plus Syndrome, although B3GLCT is not in their known target network. Thus, biological phenotypes driven by B3GLCT may be driven by the target networks of miRs that regulate this enzyme, providing additional evidence for the miRNA proxy hypothesis.
微小 RNA（miRNAs，miRs）精细地调节蛋白质表达和数以百计到数千个基因的靶标网络，这些基因控制着特定的生物学过程。它们是糖基化的关键调节剂，糖基化是最具多样性和丰富性的翻译后修饰之一。在最近的研究中，miRs 已被证明可以预测糖基化酶的生物学功能，从而提出了“miRNA 代理假说”，即“如果 miR 驱动特定的生物学表型……，那么 miR 的靶标将驱动相同的生物学表型。” 由于我们对 miR 靶标的知识不足，这个强大假说的验证受到了阻碍。靶标预测的准确性较低，假阳性率较高。在此，我们开发了一种高通量实验平台来分析 miR-靶标相互作用，即 miRFluR。我们利用该系统分析了整个人类 miRome 与β-3-葡萄糖基转移酶（B3GLCT）的相互作用，B3GLCT 是一种糖基化酶，其缺失是先天性疾病 Peters' Plus 综合征的基础。尽管该酶被多种算法预测为高度受 miR 靶向，但我们仅鉴定出 27 个下调 B3GLCT 的 miR，预测的假阳性率超过 96%。这些经验证的 miR 的功能富集分析预测了与 Peters' Plus 综合征相关的表型，尽管 B3GLCT 不在其已知的靶标网络中。因此，由 B3GLCT 驱动的生物学表型可能由调节该酶的 miR 的靶标网络驱动，为 miRNA 代理假说提供了额外的证据。
