Poster abstracts
Poster number 78 submitted by Bojing Zhu
Integrated Profiling of tRNA Modifications in the Cystic Fibrosis Cell Model with PTC suspression
Bojing Zhu (Department of Chemistry, University of Michigan, Ann Arbor), Lianwu Fu (Department of Biochemistry and Molecular Genetics, Heersink School of Medicine, The University of Alabama at Birmingham), David M Bedwell (Department of Biochemistry and Molecular Genetics, Heersink School of Medicine, The University of Alabama at Birmingham), Miten Jain (Department of Bioengineering, Northeastern University), Kristin S Koutmou (Department of Chemistry, University of Michigan, Ann Arbor)
Abstract:
Cystic fibrosis (CF) is a life-threatening genetic disorder caused by mutations in CFTR gene. Approximately 10% of patients have nonsense mutations that introduce premature termination codons (PTCs), rendering them unresponsive to current therapeutic strategies. Evidence indicates that tRNA modifications can affect suppression of PTCs and play a role in translational readthrough. In this study, we employed both direct nanopore RNA sequencing and LC-MS/MS nucleoside analysis on the total tRNA pool, derived from human bronchial epithelial cells (16HBE14o-)—including wild-type (WT), the canonical CF mutant G542X, and G542X mutants treated with various CFTR PTC readthrough-promoting activators. This integrated approach enabled the identification and quantification of 50 modifications on tRNA, as well as their sites, allowing a comprehensive mapping of the tRNA modification landscape in CF models. Our results revealed that D, m²G, m⁵C, m¹G, psudouridine, and m⁶A were highly abundant. Furthermore, modifications located within the anticodon and T loops were predominant, suggesting their crucial roles in translation fidelity and maintaining tRNA structural stability. This landscape provides insights into how tRNA modifications may modulate PTC readthrough and may help develop novel tRNA-based therapeutics for underserved populations.
Keywords: tRNA modification, cystic fibrosis, nanopore, LC-MSMS nucleoside