The RNA Methyltransferase Complex of WTAP, METTL3, and METTL14 Regulates Mitotic Clonal Expansion in Adipogenesis.

Masatoshi Kobayashi,Motoharu Awazawa,Tatsuhiko Kodama,Takashi Kadowaki,Yukiko Okazaki,Mitsuru Ohsugi,Keiko Horiuchi,Takao Hamakubo,Hironori Waki,Toshihiro Umehara,Seiichiro Aoe,Kazuyuki Tobe,Naoto Kubota,Aya Iwane,Takayoshi Sasako,Ryo Suzuki,Kohjiro Ueki,Naoki Kobayashi

doi:10.1128/mcb.00116-18

Abstract

Adipocyte differentiation is regulated by various mechanisms, of which mitotic clonal expansion (MCE) is a key step. Although this process is known to be regulated by cell cycle modulators, the precise mechanism remains unclear. N6-Methyladenosine (m6A) posttranscriptional RNA modification, whose methylation and demethylation are performed by respective enzyme molecules, has recently been suggested to be involved in the regulation of adipogenesis. Here, we show that an RNA N6-adenosine methyltransferase complex consisting of Wilms' tumor 1-associating protein (WTAP), methyltransferase like 3 (METTL3), and METTL14 positively controls adipogenesis by promoting cell cycle transition in MCE during adipogenesis. WTAP, coupled with METTL3 and METTL14, is increased and distributed in nucleus by the induction of adipogenesis dependently on RNA in vitro Knockdown of each of these three proteins leads to cell cycle arrest and impaired adipogenesis associated with suppression of cyclin A2 upregulation during MCE, whose knockdown also impairs adipogenesis. Consistent with this, Wtap heterozygous knockout mice are protected from diet-induced obesity with smaller size and number of adipocytes, leading to improved insulin sensitivity. These data provide a mechanism for adipogenesis through the WTAP-METTL3-METTL14 complex and a potential strategy for treatment of obesity and associated disorders.

Highlights

Adipocyte differentiation is regulated by various mechanisms, of which mitotic clonal expansion (MCE) is a key step
In order to investigate whether the WMM complex has the opposite regulatory function of fat mass and obesity-associated protein (FTO) in adipocyte differentiation, we first knocked down Wilms’ tumor 1-associating protein (WTAP) in 3T3-L1 cells with adeno-shWTAP and induced them to differentiate into adipocytes with standard dexamethasone–3isobutyl-1-methylxanthine (IBMX)–insulin (DMI) (Fig. 1A and B)
We observed that WTAP was constantly expressed before the induction with DMI and slightly upregulated during adipocyte differentiation at both mRNA and protein levels in embryonic fibroblasts from WT mice (MEF-WT), whereas these expression levels in MEF-Wtapϩ/Ϫ were constantly maintained at lower levels (Fig. 1E and F)

Summary

Introduction

Adipocyte differentiation is regulated by various mechanisms, of which mitotic clonal expansion (MCE) is a key step. We show that an RNA N6adenosine methyltransferase complex consisting of Wilms’ tumor 1-associating protein (WTAP), methyltransferase like 3 (METTL3), and METTL14 positively controls adipogenesis by promoting cell cycle transition in MCE during adipogenesis. Consistent with this, Wtap heterozygous knockout mice are protected from diet-induced obesity with smaller size and number of adipocytes, leading to improved insulin sensitivity These data provide a mechanism for adipogenesis through the WTAP-METTL3-METTL14 complex and a potential strategy for treatment of obesity and associated disorders. Increasing numbers of studies have recently demonstrated that WTAP conforms in a complex in the nucleus with METTL3 and METTL14, which methylate N6-adenosines of RNA in mammalian cells, involving posttranscriptional mRNA metabolism, splicing, and cell cycle regulation [7,8,9,10]

Methods

Results

Conclusion