Epitranscriptomic regulation by m6A RNA methylation in brain development and diseases

Abstract

Cellular RNAs are pervasively tagged with diverse chemical moieties, collectively called epitranscriptomic modifications. The methylation of adenosine at N⁶ position generates N⁶-methyladenosine (m⁶A), which is the most abundant and reversible epitranscriptomic modification in mammals. The m⁶A signaling is mediated by a dedicated set of proteins comprised of writers, erasers, and readers. Contrary to the activation-repression binary view of gene regulation, emerging evidence suggests that the m⁶A methylation controls multiple aspects of mRNA metabolism, such as splicing, export, stability, translation, and degradation, culminating in the fine-tuning of gene expression. Brain shows the highest abundance of m⁶A methylation in the body, which is developmentally altered. Within the brain, m⁶A methylation is biased toward neuronal transcripts and sensitive to neuronal activity. In a healthy brain, m⁶A maintains several developmental and physiological processes such as neurogenesis, axonal growth, synaptic plasticity, circadian rhythm, cognitive function, and stress response. The m⁶A imbalance contributes to the pathogenesis of acute and chronic CNS insults, brain cancer, and neuropsychiatric disorders. This review discussed the molecular mechanisms of m⁶A regulation and its implication in the developmental, physiological, and pathological processes of the brain.

Keywords: Brain development; N6-methyladenosine; brain physiology; neurological disorders; post-transcriptional regulation.

Figure 1.

Major epitranscriptomic modifications. Mature RNAs can undergo 172 types of chemical modifications; 72 of those are variants of methyl group modifications. The most studied epitranscriptomic modifications are N⁶-methyladenosine (m⁶A), pseudouridine (ψ), 5-methylcytidine (m⁵C), N¹-methyladenosine (m¹A), ribose 2'-O-methylation (Nm), and N⁴-acetylcytidine (ac4C).

Figure 2.

Molecular machinery of m⁶A methylation. The m⁶A modification predominantly occurs at consensus RRACH (R is a purine base, A is m⁶A-modified adenine, and H is a non-guanine base) motif enriched in the 3'-UTR of mRNAs. The m⁶A is deposited by a multi-subunit methylase complex (aka writer complex) primarily composed of a catalytic subunit METTL3 and a regulatory subunit METTL14. The m⁶A methylation is reversible and the methylated RNAs are demethylated by FTO and ALKBH5, with distinct erasure mechanisms. The m⁶A readers bind the methyl group through a characteristic YTH domain to propagate the m⁶A signaling. Additionally, the m⁶A methylation is repelled by m⁶A anti-readers G3BP1/2 that preferentially bind to unmethylated m⁶A motifs in the RNA. WTAP: Wilms’ tumor-associated protein; METTL3: methyltransferase like-3; METTL14: methyltransferase like-14; FTO: fat mass and obesity-associated; YTH: YT521-B homology; G3BP1/2: GTPase-activating protein (SH3 domain) binding protein 1 and 2.

Figure 3.

Techniques to study m⁶A methylation. LC-MS/MS can quantitate the m⁶A modification (a). MeRIP-seq can identify the transcriptome-wide m⁶A profiles. MAZTER-seq can show the m⁶A sites along with the stoichiometry of methylation with single-nucleotide resolution (b). CRISPR/Cas9 genomic editing using METTL3 or FTO fused to inactive dCas9 targeted near the vicinity of m⁶A site by sequence-specific gRNA can modulate site-specific m⁶A methylation (c).

Figure 4.

Post-transcriptional regulation by m⁶A methylation. The m⁶A methylation occurs co-transcriptionally in the nascent pre-mRNAs. The fate of methylated transcripts is dictated by m⁶A readers that bind and recruit diverse cellular machinery. In the nucleus, YTHDC1 binds to exonic m⁶A and recruits splicing regulatory factor SRSF3 to promote exon inclusion and thereby controls alternate splicing (a). The m⁶A writer facilitates the nuclear export of spliced mRNAs by interacting with export machinery TREX complex to guide them through the nuclear pore complex to the cytoplasm (b). In the cytoplasm, the presence of m⁶A repels the binding of stress granule proteins G3BP1/2 that regulates the stability of target RNA (c). Binding of YTHDF1 to methylated RNA recruits translation machinery leading to RNA translation (d). In contrast, YTHDF2 recruits mRNA decay enzymes in the P-bodies leading to degradation of the methylated transcripts (e). SRSF3: serine/arginine-rich splicing factor 3; YTHDC1: YT521-B homology domain-containing protein 1; G3BP1/2: GTPase activating protein (SH3 domain) binding protein 1/2; YTHDF1: YT521-B homology domain-containing family protein 1; YTHDF2: YT521-B homology domain-containing family protein 2; NPC: nuclear pore complex.

Figure 5.

Role of m⁶A methylation in brain development, physiology, and disease. The m⁶A, with its multifaceted role in mRNA processing, is considered as the fifth nucleobase in the mammalian brain. In the developing brain, m⁶A gradually increases to control axonogenesis, neurogenesis, and gliogenesis. In the adult brain, m⁶A maintains essential physiological processes such as synaptic plasticity, cognitive function, and circadian rhythm. The m⁶A dysregulation is implicated in many pathological processes, including chronic neurodegeneration, acute brain injury, and tumorigenesis. METTL3: methyltransferase like-3; METTL14: methyltransferase like-14; WTAP: Wilms’ tumor-associated protein; FTO: fat mass and obesity-associated; ALKBH5: alkB homolog 5; YTH: YT521-B homology.