Poster Presentation 43rd Lorne Genome Conference 2022

Detection of m6A and m5C RNA modifications at single-molecule resolution using Nanopore sequencing (#117)

Pablo Acera 1 2 , Marco Guarnacci 1 2 , Aditya Sethi 1 2 , Akansha Ravindran 1 2 , Jiajia Xu 1 , Katrina Woodward 1 , William Hamilton 3 , Shaam Al Abed 4 , Rippei Hayashi 1 , Thomas Preiss 1 2 , Vihandha Wickramasinghe 3 , Nathalie Dehorter 4 , Nikolay Shirokikh 1 , Eduardo Eyras 1 2 5 6
  1. Cancer and Genome Sciences, The John Curtin School of Medical Research, Australian National University, Canberra, ACT, Australia
  2. EMBL Australia Partner Laboratory Network at the Australian National University, Canberra, ACT, Australia
  3. Peter MacCallum Cancer Centre, Melbourne, Victoria, Australia
  4. Neuroscience, The John Curtin School of Medical Research, Australian National University, Canberra, ACT, Australia
  5. Hospital del Mar Medical Research Institute (IMIM), Barcelona, Catalonia, Spain
  6. Catalan Institution for Research and Advanced Studies (ICREA), Barcelona, Catalonia, Spain

The expanding field of epitranscriptomics has been setting to rival the epigenome in the diversity of biological process involvement, prominently linking biochemical modifications of the RNA to development and disease onset. However, the identification of modifications in individual RNA molecules remains challenging. Here we describe CHEUI, a new computational approach to identify N6-methyladenosine (m6A) and 5-methylcytidine (m5C) using signals from Nanopore direct RNA sequencing reads at single-nucleotide and single-molecule resolution. CHEUI uses a two-stage neural network to accurately predict methylation in individual reads and transcriptomic sites in a single condition, as well as differential methylation between any two conditions. Using extensive benchmarking with Nanopore data derived from in vitro modified and non-modified transcripts as well as cells with or without methylation enzymes, CHEUI showed higher accuracy than other existing methods in the prediction of m6A and m5C sites and their stoichiometry levels, while maintaining a lower number of false positives. CHEUI’s ability to detect RNA modifications can be expanded to other modifications to unveil the full span of the epitranscriptome in normal and disease conditions.