Mulroney Logan, Taylor Henry J, Lee Angela, Swift Amy J, Zdravkov Mihail, Bonnycastle Lori L, Brooks Shelise Y, Lee Brian N, Fitzgerald Tomas, Narisu Narisu, Biesecker Leslie G, Erdos Michael R, Nicassio Francesco, Birney Ewan, Collins Francis S, Taylor D Leland
bioRxiv. 2025 Jun 12:2025.06.12.659352. doi: 10.1101/2025.06.12.659352.
RNA modifications are critical regulators of gene expression and cellular processes; however, the epitranscriptome is less well studied than the epigenome. Here, we studied transcriptome-wide changes in RNA modifications and expression levels in two human pancreatic beta-cell lines, EndoC-BH1 and EndoC-BH3, after one hour of glucose stimulation. Using direct RNA nanopore sequencing (dRNA-seq), we measured N6-methyladenosine (m6A), 5-methylcytosine (m5C), inosine, and pseudouridine concurrently across the transcriptome. We developed a differential RNA modification method and identified 1,697 differentially modified sites (DMSs) across all modifications. These DMSs were largely independent of changes in gene expression levels and enriched in transcripts for type 2 diabetes (T2D) genes. Our study demonstrates how dRNA-seq can be used to detect and quantify RNA modification changes in response to cellular stimuli at the single-nucleotide level and provides new insights into RNA-mediated mechanisms that may contribute to normal beta-cell response and potential dysfunction in T2D.
RNA修饰是基因表达和细胞过程的关键调节因子；然而，与表观基因组相比，对表转录组的研究较少。在此，我们研究了两种人类胰腺β细胞系EndoC-BH1和EndoC-BH3在葡萄糖刺激1小时后RNA修饰和表达水平的全转录组变化。使用直接RNA纳米孔测序（dRNA-seq），我们在转录组范围内同时测量了N6-甲基腺苷（m6A）、5-甲基胞嘧啶（m5C）、次黄嘌呤和假尿苷。我们开发了一种差异RNA修饰方法，并在所有修饰中鉴定出1697个差异修饰位点（DMS）。这些DMS在很大程度上与基因表达水平的变化无关，并且在2型糖尿病（T2D）基因的转录本中富集。我们的研究展示了dRNA-seq如何用于在单核苷酸水平上检测和量化响应细胞刺激的RNA修饰变化，并为可能有助于正常β细胞反应和T2D潜在功能障碍的RNA介导机制提供了新见解。
