Deficiency of N1-Adenine Methyltransferase Aggravates RNA and Protein Aggregation.

RNA modifications regulate diverse aspects of transcripts' function and stability. Among these, N1-methyladenine (mA) is a reversible mark primarily installed by the TRMT6/TRMT61A methyltransferase on tRNA, though it is also found on other RNA types. mA has been implicated in protecting mRNAs during acute protein misfolding stress. However, the role of mA under chronic proteotoxic conditions, such as intracellular amyloid aggregation, remains poorly understood. To address this gap, we examined the effects of reduced N1-adenine methylation in human cells undergoing amyloidogenesis. Suppression of the methyltransferase TRMT61A or overexpression of the mA-specific demethylase ALKBH3 enhanced amyloid aggregation. A deficiency of N1-adenine methylation also impaired the expression of a reporter mRNA-encoded protein, highlighting the protective role of mA in safeguarding transcript functionality. Proteomic analysis of amyloid aggregates from TRMT61A-deficient cells revealed increased co-aggregation of bystander proteins, particularly those with known RNA-binding activity. At the same time, the aggregates from methylation-deficient cells contained elevated levels of mRNAs. Collectively, our findings support a role for mA in preventing RNA entanglement within aggregates and limiting RNA-mediated propagation of protein co-aggregation.