FTO-dependent demethylation of N6-methyladenosine regulates mRNA splicing and is required for adipogenesis

Abstract

The role of Fat Mass and Obesity-associated protein (FTO) and its substrate N6-methyladenosine (m6A) in mRNA processing and adipogenesis remains largely unknown. We show that FTO expression and m6A levels are inversely correlated during adipogenesis. FTO depletion blocks differentiation and only catalytically active FTO restores adipogenesis. Transcriptome analyses in combination with m6A-seq revealed that gene expression and mRNA splicing of grouped genes are regulated by FTO. M6A is enriched in exonic regions flanking 5′- and 3′-splice sites, spatially overlapping with mRNA splicing regulatory serine/arginine-rich (SR) protein exonic splicing enhancer binding regions. Enhanced levels of m6A in response to FTO depletion promotes the RNA binding ability of SRSF2 protein, leading to increased inclusion of target exons. FTO controls exonic splicing of adipogenic regulatory factor RUNX1T1 by regulating m6A levels around splice sites and thereby modulates differentiation. These findings provide compelling evidence that FTO-dependent m6A demethylation functions as a novel regulatory mechanism of RNA processing and plays a critical role in the regulation of adipogenesis.

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].…”

Nuclear TARBP2 drives oncogenic dysregulation of RNA splicing and decay

“…The prevalent internal RNA modification mark N(6)-methyladenosine (m 6 A) has been reported to play a role in regulating most facets of the RNA life cycle, including regulation of pre-mRNA splicing, mRNA stability, and mRNA translation (Lin et al, 2016;Liu et al, 2015;Wang et al, 2015Wang et al, , 2014Xiao et al, 2016;Zhao et al, 2014). Recently, work by us (Alarcón et al, 2015) and others (Ke et al, 2017;Knuckles et al, 2017) has established that m 6 A marks are deposited in the nucleus and are proposed to function in many nuclear regulatory processes, including microRNA and messenger RNA processing.…”

Kaposi's Sarcoma-Associated Herpesvirus Utilizes and Manipulates RNA N 6 -Adenosine Methylation To Promote Lytic Replication

“…Three methyltransferases, including methyltransferase-like 3 (METTL3), methyltransferase-like 14 (METTL14), and Wilms' tumor 1-associated protein (WTAP), act as m 6 A writers and catalyze RNA m 6 A at specific sites with the consensus sequence [(G/A)GAC, where the underlined adenosine is the methylation site] (11,12). Two demethylases, fat mass-and obesity-associated protein (FTO) and AlkB homolog 5 (ALKBH5), both of which act as m 6 A erasers, reverse this process (13)(14)(15)(16)(17). Most m 6 A sites are located near the transcription start sites, exonic regions flanking splicing sites, stop codons, and the 3= untranslated region (3= UTR) (1, rabbit anti-m 6 A antibody and control IgG, followed by conversion of the input RNAs and the RNA immunoprecipitation (RIP) products into cDNAs with reverse transcriptase (RT) (Fig.…”