RNA-Methylation-Dependent RNA Processing Controls the Speed of the Circadian Clock

Abstract

The eukaryotic biological clock involves a negative transcription-translation feedback loop in which clock genes regulate their own transcription and that of output genes of metabolic significance. While around 10% of the liver transcriptome is rhythmic, only about a fifth is driven by de novo transcription, indicating mRNA processing is a major circadian component. Here, we report that inhibition of transmethylation reactions elongates the circadian period. RNA sequencing then reveals methylation inhibition causes widespread changes in the transcription of the RNA processing machinery, associated with m(6)A-RNA methylation. We identify m(6)A sites on many clock gene transcripts and show that specific inhibition of m(6)A methylation by silencing of the m(6)A methylase Mettl3 is sufficient to elicit circadian period elongation and RNA processing delay. Analysis of the circadian nucleocytoplasmic distribution of clock genes Per2 and Arntl then revealed an uncoupling between steady-state pre-mRNA and cytoplasmic mRNA rhythms when m(6)A methylation is inhibited.

Crystal structure of the YTH domain of YTHDF2 reveals mechanism for recognition of N6-methyladenosine

“…The recent discoveries of RNA m 6 A methyltransferase complex METTL3/METTL14/WTAP and demethylases FTO and ALKBH5 prove the reversibility of m6A modification [2][3][4][5][6]. This modification plays important roles in various biological processes, including circadian rhythms [7], RNA splicing [8], yeast meiosis [9], and embryonic stem cell self-renewal [10]. Two recent studies show that YTH domain family 2 (YTHDF2) and other YTHDF proteins preferentially bind to m 6 A-containing mRNA in vivo and in vitro and regulate localization and stability of the bound mRNA [8,11].…”

YTHDF3 facilitates translation and decay of N6-methyladenosine-modified RNA

“…m 6 A is dynamically deposited, removed, and recognized by m 6 A methyltransferases ("writers"), demethylases ("erasers"), and m 6 A-specific binding proteins ("readers"), respectively. The "writer" METTL3 is crucial in regulating stem cell pluripotency, cell differentiation, and circadian period [6][7][8], whereas depletion of the "erasers" FTO and ALKBH5 has revealed their roles in energy homeostasis, adipocyte differentiation, and fertility in mice [9][10][11].…”

Identification of factors required for m6A mRNA methylation in Arabidopsis reveals a role for the conserved E3 ubiquitin ligase HAKAI

“…N 6‐methyladenosine (m 6 A) is the most prevalent internal mRNA modification found in eukaryotes, and has received a burst of interest in recent years (Meyer & Jaffrey, 2014; Fray & Simpson, 2015; Yue et al ., 2015). m 6 A appears to be involved in a broad range of biological processes including mRNA export from the nucleus (Fustin et al ., 2013), regulation of splicing (Alarcón et al ., 2015b; Haussmann et al ., 2016; Lence et al ., 2016), mRNA translatability and stability (Wang et al ., 2014a,b, 2015; Bodi et al ., 2015; Zhou et al ., 2015), alternative polyadenylation site choice (Ke et al ., 2015) and other mechanisms accompanying RNA maturation (Meyer & Jaffrey, 2014; Yue et al ., 2015). m 6 A is essential for the earliest stages of pattern formation in plants (Zhong et al ., 2008; Bodi et al ., 2012; Shen et al ., 2016) and metazoans (Meyer & Jaffrey, 2014; Geula et al ., 2015; Yue et al ., 2015; Haussmann et al ., 2016; Lence et al ., 2016), linked with diseases in humans and other mammalian species (Jia et al ., 2011; Zheng et al ., 2013) and is required for meiosis in Saccharomyces cerevisiae (Clancy et al ., 2002).…”

“…m 6 A is essential for the earliest stages of pattern formation in plants (Zhong et al ., 2008; Bodi et al ., 2012; Shen et al ., 2016) and metazoans (Meyer & Jaffrey, 2014; Geula et al ., 2015; Yue et al ., 2015; Haussmann et al ., 2016; Lence et al ., 2016), linked with diseases in humans and other mammalian species (Jia et al ., 2011; Zheng et al ., 2013) and is required for meiosis in Saccharomyces cerevisiae (Clancy et al ., 2002). Reduced levels of m 6 A also affect circadian period (Fustin et al ., 2013) and are critical for stem cell differentiation in mammals (Geula et al ., 2015). …”