Deficiency of N1-Adenine Methyltransferase Aggravates RNA and Protein Aggregation

RNA modifications regulate diverse aspects of transcripts' function and stability. Among these, N1-methyladenine (m¹A) is a reversible mark primarily installed by the TRMT6/TRMT61A methyltransferase on tRNA, though it is also found on other RNA types. m¹A has been implicated in protecting mRNAs during acute protein misfolding stress. However, the role of m¹A 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 m¹A-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 m¹A 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 m¹A in preventing RNA entanglement within aggregates and limiting RNA-mediated propagation of protein co-aggregation.