Abstract
We previously showed that the N6-methyladenosine (m6A) mRNA methylase is essential during Arabidopsis thaliana embryonic development. We also demonstrated that this modification is present at varying levels in all mature tissues. However, the requirement for the m6A in the mature plant was not tested. Here we show that a 90% reduction in m6A levels during later growth stages gives rise to plants with altered growth patterns and reduced apical dominance. The flowers of these plants commonly show defects in their floral organ number, size, and identity. The global analysis of gene expression from reduced m6A plants show that a significant number of down-regulated genes are involved in transport, or targeted transport, and most of the up-regulated genes are involved in stress and stimulus response processes. An analysis of m6A distribution in fragmented mRNA suggests that the m6A is predominantly positioned toward the 3′ end of transcripts in a region 100–150 bp before the poly(A) tail. In addition to the analysis of the phenotypic changes in the low methylation Arabidopsis plants we will review the latest advances in the field of mRNA internal methylation
Highlights
N 6-methyladenosine (m6A) is a ubiquitous base modification found internally in the mRNA of most Eukaryotes
We previously showed that the N6-methyladenosine (m6A) mRNA methylase is essential during Arabidopsis thaliana embryonic development
The role of mRNA methylation in normal mRNA metabolism is not known, and it remains possible that methylation plays a positive or negative role in post-transcriptional gene silencing; for this reason we wished to avoid an RNAi based approach for reducing MTA levels
Summary
N 6-methyladenosine (m6A) is a ubiquitous base modification found internally in the mRNA of most Eukaryotes. Whilst other types of post-transcriptional base modification, such as C-to-U editing, or A-to-I conversions result in a change following reverse transcription, m6A is recognized as adenosine and so is not revealed by cDNA sequencing For this reason it is technically demanding to map its position in specific messages, and this has only been achieved in two, relatively abundant, transcripts. In Rous sarcoma virus a 1865 nt region of the genomic RNA contains seven m6A sites in a GAC context (Kane and Beemon, 1985; Csepany et al, 1990), and in bovine prolactin mRNA, methylation occurs at a single AGACU site within the 3 untranslated region (UTR; Horowitz et al, 1984) In both cases, this conformed to an extended consensus sequence RRACH (where R represents purine, A is the methylation site, and H is any base other than G), previously proposed by Schibler et al (1977) Consistent with this, the partially purified mammalian mRNA methylase shows strongest activity for GGACU and, to a lesser extent, AGACU sequences (Harper et al, 1990). The position of m6A in the meiotic kinase IME2, was mapped to the 3 half of the message, a region where the translational suppressor Khd has previously been predicted to bind (Hasegawa et al, 2009; Bodi et al, 2010)
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