Sakhawala Rima M, Tirgar Reyhaneh, Vieux Karl-Frédéric, Haskell Dustin, Yu Guoyun, Zinovyeva Anna, McJunkin Katherine
Laboratory of Cellular and Developmental Biology, National Institute of Diabetes and Digestive and Kidney Diseases Intramural Research Program, National Institutes of Health, Bethesda, Maryland, USA.
Department of Biology, Johns Hopkins University, Baltimore, Maryland, USA.
bioRxiv. 2025 Jun 2:2025.04.11.648421. doi: 10.1101/2025.04.11.648421.
MicroRNAs (miRNAs) are short RNAs that post-transcriptionally regulate gene expression. In canonical miRNA biogenesis, primary miRNAs are transcribed from intergenic loci or intronic regions by RNA polymerase II, sequentially cleaved by the Microprocessor complex and Dicer, and resulting mature miRNAs are loaded into Argonaute to repress target mRNAs. A minority of miRNAs are generated via noncanonical biogenesis pathways that bypass the Microprocessor and/or Dicer. Here, we describe a new Pol III-dependent, Microprocessor-independent, and Dicer-dependent biogenesis pathway exemplified by the family in . Although the family loci reside in intronic regions of protein-coding genes, we show that the miRNAs are derived from independent Pol III transcripts. Unlike other Pol III-dependent miRNAs, the family small RNAs are the dominant species derived from their loci, rather than fragments of a larger functional noncoding RNA. These germline-enriched miRNAs are loaded in multiple miRNA Argonautes, including the recently-characterized germline Argonaute ALG-5, which we demonstrate is repressive when tethered to a reporter transcript. We extend these findings, identifying additional Pol III-transcribed and noncanonical small RNAs in and human datasets, including human miR-4521. These young, noncanonical miRNAs may represent an early snapshot in the evolution of de novo miRNA genes.
微小RNA（miRNA）是一类在转录后水平调控基因表达的短RNA。在经典的miRNA生物合成过程中，初级miRNA由RNA聚合酶II从基因间区域或内含子区域转录而来，先后被微处理器复合体和Dicer切割，产生的成熟miRNA被装载到AGO蛋白中以抑制靶标mRNA。少数miRNA通过绕过微处理器和/或Dicer的非经典生物合成途径产生。在此，我们描述了一种新的由RNA聚合酶III依赖、不依赖微处理器且依赖Dicer的生物合成途径，以秀丽隐杆线虫中的miRNA家族为例。尽管该miRNA家族基因座位于蛋白质编码基因的内含子区域，但我们发现这些miRNA来源于独立的RNA聚合酶III转录本。与其他RNA聚合酶III依赖的miRNA不同，该家族的小RNA是其基因座产生的主要产物，而非较大功能性非编码RNA的片段。这些在生殖系中富集的miRNA被装载到多种miRNA AGO蛋白中，包括最近鉴定的生殖系AGO蛋白ALG-5，我们证明当它与报告转录本结合时具有抑制作用。我们扩展了这些发现，在秀丽隐杆线虫和人类数据集中鉴定出了其他由RNA聚合酶III转录的非经典小RNA，包括人类miR-4521。这些年轻的非经典miRNA可能代表了新生miRNA基因进化过程中的早期状态。
