The epitranscriptomic m6A RNA modification modulates synaptic function in ageing and in a mouse model of synucleinopathy

Abstract

N6-methyladenosine (m6A) is the most abundant and conserved transcriptional modification in eukaryotic RNA, regulating RNA fate. While the functions of m6A in the development of the mammalian brain have been extensively studied, its roles in synaptic plasticity, cognitive decline, motor function, or other brain circuits remain underexplored. To date, the role of this modification in Parkinsons disease (PD) and other synucleinopathies has been largely unknown. Here, we investigated the m6A epitranscriptome in a mouse model of synucleinopathy. We performed m6A RNA immunoprecipitation sequencing (meRIP-seq) to obtain the m6A epitranscriptome of the midbrain in young (3 mo) and aged (15 mo) A30P-aSyn transgenic mice (aSyn Tg) and C57BL6 control wild type (Wt) mice. We observed hypermethylation of synaptic genes in 3 mo aSyn Tg mice compared to age-matched Wt mice. This methylation was reduced during ageing, with synaptic genes becoming increasingly hypomethylated. Using immunofluorescence imaging alongside biochemical analysis, we further investigated the expression of m6A regulatory enzymes -- writer, N6-Adenosine-Methyltransferase Complex Catalytic Subunit (METTL3); reader, YTH N6-methyladenosine RNA-binding protein (YTHDF1); and eraser, fat mass and obesity-associated protein (FTO) -- in the cortex, striatum, hippocampus, and cerebellum of Wt and aSyn Tg mice, as well as in primary cortical neuronal cultures. We observed that the levels of METTL3, YTHDF1 and FTO were similar between Wt and aSyn Tg mice. Interestingly, the writer protein METTL3 was found in both the nucleus and in the post-synaptic compartment in neuronal cultures. Our findings suggest that alterations in the regulation of m6A RNA methylation may be associated with neurodegeneration and ageing and that this level of epitranscriptomic regulation plays a significant role at the synapse.