Abstract B041: Identification of m3C modification sites within mRNAs of pancreatic cancer cells by long-read direct RNA sequencing

Abstract

Abstract Chemical modification is a precise and effective way to control the biogenesis, stability, and function of biological molecules, such as nucleic acids, proteins, and lipids. More than 170 different types of modifications have been discovered in coding and non-coding RNAs, constituting a diverse RNA modification landscape, called epitranscriptome. RNA modifications were mediated by three types of proteins: writers, erasers, and readers. Recently, RNA modification machinery has been dysregulated in several types of cancers and may be promising targets for cancer therapy. 3-methylcytidine (m3C), which is catalyzed by METTL2A, 2B, 6, and 8, is found at position 32 of cytoplasmic tRNAs in organisms ranging from yeast to humans, however, their functions and regulatory machinery in mRNA and non-coding RNA has not been clarified because there were no effective anti- m3C antibodies. Here we discovered that the expression of m3C writers was higher in pancreatic tumors than in noncancerous tissues. We further found that METTL2A expression is associated with a poor prognosis in pancreatic cancer patients. We also showed that METTL2A depletion led to a reduction in the growth of AsPC1 cells. We next attempted to identify the m3C sites in cellular transcripts regulated by METTL2A in AsPC1 cells using long-read direct RNA sequencing. We corrected the transcriptome using ESPRESSO and searched for the RNA modifications using Nanocompore. We then found that METTL2A targets the NYCYN motif in mRNAs and mitochondrial rRNAs. Moreover, we found that transcripts with METTL2A-mediated m3C modification were upregulated in AsPC-1 cells knocked down by METTL2A. Our study provides evidence for METTL2A-mediated down-regulation of target mRNAs via m3C modification in the survival of pancreatic tumor cells. Citation Format: Shuhei Mitsutomi, Anzu Sugawara, Kenji Takahashi, Yusuke Mizukami, Nobuyoshi Akimitsu, Kenzui Taniue. Identification of m3C modification sites within mRNAs of pancreatic cancer cells by long-read direct RNA sequencing [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Cancer Evolution and Data Science: The Next Frontier; 2023 Dec 3-6; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_2):Abstract nr B041.