and Abstract ! The RNA methylome, in contrast to the DNA methylome, has barely been explored in the context of normal and diseased brain development and function. RNA cytosine methylation (m5C), characterized by the addition of a methyl group on the 5' position of a cytosine in RNA, is a novel modification only recently characterized in mammalian tissue and most abundant in tRNAs. NSUN2, a tRNA methyltransferase that methylates the variable loop of tRNA, is expressed at high levels in the central nervous system and has been linked to neurodevelopmental defects in humans and mice. Humans with a mutation in the NSUN2 gene exhibit intellectual disability accompanied by other neurological abnormalities, including microcephaly, speech delays, and other cognitive impairments. Likewise, adult NSUN2 knockout mice exhibit deficits in cognitive and emotional behavior, a phenotype which may be related to altered methylation at distinct tRNA cytosines during embryonic stages of brain development leading to impaired neuronal survival and synaptic puncta in the cortex. However, it is still unknown whether tRNA methylation patterns regulate brain function or behavioral outcomes outside the realm of development. In the current proposal, I will focus on the regulation of RNA cytosine methylation in the mouse brain and its role in cognitive behavior.
In Aim 1 we will use viral overexpression and knockdown of NSUN2 in the adult mouse PFC to elucidate the effect of altered NSUN2 expression on tRNA methylation in the PFC.
Aim 2 will investigate cognitive and anxiety-like behaviors of NSUN2-overexpressing and knockdown mice.
Aim 3 will identify the downstream molecular consequences of tRNA methylation on the accumulation of tRNA fragments and presence of stress granules and processing bodies. The proposed research is significant because RNA cytosine methylation has still not been explored in the adult brain, and understanding the role of tRNA methylation in the adult brain may elucidate mechanisms underlying aberrant brain function during adulthood.
and Public Health Relevance ! The proposed research uses an animal model to focus on the downstream molecular and behavioral effects of tRNA methylation in the adult brain. Previous work shows impairments in cognition for both humans and animals with altered NSUN2 expression, which plays a crucial role in tRNA methylation. Here we will use viral overexpression and knockdown of NSUN2 in the adult prefrontal cortex to elucidate the effect of NSUN2- induced tRNA methylation on stress-related molecular markers and behavior. This research will enhance our understanding of how the RNA methylome can contribute to brain function during adulthood.
