Uncovering the Epitranscriptome: A Review on mRNA Modifications and Emerging Frontiers

Background/Objectives: Messenger RNA (mRNA) modifications regulate key steps in gene expression, including splicing, translation, and stability. Despite over 300 known RNA modifications, the relatively small subset occurring in mRNA remains understudied compared with tRNA and rRNA. This review aims to systematically evaluate 15 known naturally occurring mRNA-specific modifications, rank them by publication frequency, and highlight emerging frontiers in epitranscriptomics, including discovering new naturally occurring mRNA modifications and environmental RNA (eRNA) epitranscriptomics. Methods: We conducted a structured literature review of PubMed-indexed publications to rank mRNA modifications by citation prevalence. Key modifications such as m6A, m5C, Ψ, and m1A were analyzed in terms of enzymatic machinery (“writers,” “erasers,” and “readers”), molecular functions, and physiological relevance. We also reviewed technological advances, with a focus on nanopore sequencing for detection of RNA modifications in native and environmental contexts. Results: The modification m6A was identified as the most studied mRNA modification, followed by Ψ, m5C, and A-to-I editing (inosine). These modifications influence diverse mRNA processes, including translation efficiency, localization, and immune evasion. Cap-specific modifications such as Cap0, Cap1, and Cap2 were also described, highlighting their role in transcript stability and innate immune regulation. Advances in nanopore sequencing have enabled direct detection of RNA modifications and offer promise for eRNA (environmental RNA) surveys. The potential for nanopore sequencing of many other of the 335 known RNA modifications in the MODOMICS database using existing nanopore technologies is also discussed. Conclusions: mRNA modifications represent a critical, yet incompletely mapped, layer of gene regulation. Continued research—especially using nanopore and machine learning technologies—will help uncover their full biological significance. Exploration of eRNA and identifying new mRNA modifications will redefine our understanding of RNA biology.