The N¹-Methyladenosine Methylome of Petunia mRNA

N¹-methyladenosine is a unique type of base methylation in that it blocks Watson-Crick base pairing and introduces a positive charge. m¹A is prevalent in yeast and mammalian mRNA and plays a functional role. However, little is known about the abundance, dynamics, and topology of this modification in plant mRNA. Dot blotting and liquid chromatography tandem mass spectrometry analyses revealed a dynamic pattern of m¹A mRNA modification in various tissues and at different developmental stages in petunia (Petunia hybrida), a model system for plant growth and development. We performed transcriptome-wide profiling of m¹A in petunia mRNA by m¹A mRNA immunoprecipitation followed by a deep-sequencing approach (m¹A-seq, using an m¹A-specific antibody). m¹A-seq analysis identified 4,993 m¹A peaks in 3,231 genes expressed in petunia corollas; there were 251 m¹A peaks in which A residues were partly replaced by thymine and/or reverse transcription stopped at an adenine site. m¹A was enriched in coding sequences, with single peaks located immediately after start codons. Ethylene treatment upregulated 400 m¹A peaks in 375 mRNAs and downregulated 603 m¹A peaks in 530 mRNAs in petunia corollas; 975 m¹A peaks in mRNA were only detected in corollas treated with air and 430 were only detected in corollas treated with ethylene. Silencing of petunia tRNA-specific methyltransferase 61A (PhTRMT61A) reduced the m¹A level in mRNA in vivo and in vitro. In addition, PhTRMT61A silencing caused abnormal leaf development, and the PhTRMT61A protein was localized to the nucleus. Thus, m¹A in mRNA is an important epitranscriptome marker and plays a role in plant growth and development.