Multiple Functions and Mechanisms Underlying the Role of METTL3 in Human Cancers.
Zheng Wenhui,Dong Xiaoshen,Zhao Yan,Wang Shuo,Jiang Haiyang,Zhang Mingdi,Zheng Xinyu,Gu Ming
Frontiers in oncology
Methyltransferase-like 3 (METTL3), a predominantly catalytic enzyme in the N-methyladenosine (mA) methyltransferase system, is dysregulated and plays a dual role (oncogene or tumor suppressor) in different human cancers. The expression and pro- or anticancer role of METTL3 in different cancers remain controversial. METTL3 is implicated in many aspects of tumor progression, including tumorigenesis, proliferation, invasion, migration, cell cycle, differentiation, and viability. Most underlying mechanisms involve multiple signaling pathways that rely on mA-dependent modification. However, METTL3 can also modulate the cancer process by directly promoting the translation of oncogenes via interaction with the translation initiation machinery through recruitment of eukaryotic translation initiation factor 3 subunit h (eIF3h). In this review, we summarized the current evidence on METTL3 in diverse human malignancies and its potential as a prognostic/ therapeutic target.
Interactions, localization, and phosphorylation of the mA generating METTL3-METTL14-WTAP complex.
Schöller Eva,Weichmann Franziska,Treiber Thomas,Ringle Sam,Treiber Nora,Flatley Andrew,Feederle Regina,Bruckmann Astrid,Meister Gunter
RNA (New York, N.Y.)
-methyladenine (mA) is found on many eukaryotic RNAs including mRNAs. mA modification has been implicated in mRNA stability and turnover, localization, or translation efficiency. A heterodimeric enzyme complex composed of METTL3 and METTL14 generates mA on mRNAs. METTL3/14 is found in the nucleus where it is localized to nuclear speckles and the splicing regulator WTAP is required for this distinct nuclear localization pattern. Although recent crystal structures revealed how the catalytic MT-A70 domains of METTL3 and METTL14 interact with each other, a more global architecture including WTAP and RNA interactions has not been reported so far. Here, we used recombinant proteins and mapped binding surfaces within the METTL3/14-WTAP complex. Furthermore, we identify nuclear localization signals and identify phosphorylation sites on the endogenous proteins. Using an in vitro methylation assay, we confirm that monomeric METTL3 is soluble and inactive while the catalytic center of METTL14 is degenerated and thus also inactive. In addition, we show that the C-terminal RGG repeats of METTL14 are required for METTL3/14 activity by contributing to RNA substrate binding. Our biochemical work identifies characteristic features of METTL3/14-WTAP and reveals novel insight into the overall architecture of this important enzyme complex.
N6-methyladenosine alters RNA structure to regulate binding of a low-complexity protein.
Liu Nian,Zhou Katherine I,Parisien Marc,Dai Qing,Diatchenko Luda,Pan Tao
Nucleic acids research
N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNA (mRNA), and affects almost every stage of the mRNA life cycle. The YTH-domain proteins can specifically recognize m6A modification to control mRNA maturation, translation and decay. m6A can also alter RNA structures to affect RNA-protein interactions in cells. Here, we show that m6A increases the accessibility of its surrounding RNA sequence to bind heterogeneous nuclear ribonucleoprotein G (HNRNPG). Furthermore, HNRNPG binds m6A-methylated RNAs through its C-terminal low-complexity region, which self-assembles into large particles in vitro. The Arg-Gly-Gly repeats within the low-complexity region are required for binding to the RNA motif exposed by m6A methylation. We identified 13,191 m6A sites in the transcriptome that regulate RNA-HNRNPG interaction and thereby alter the expression and alternative splicing pattern of target mRNAs. Low-complexity regions are pervasive among mRNA binding proteins. Our results show that m6A-dependent RNA structural alterations can promote direct binding of m6A-modified RNAs to low-complexity regions in RNA binding proteins.
Role of N-methyladenosine modification in cancer.
Deng Xiaolan,Su Rui,Feng Xuesong,Wei Minjie,Chen Jianjun
Current opinion in genetics & development
As the most abundant internal modification in eukaryotic messenger RNAs identified, N-methyladenosine (mA) has been shown recently to play essential roles in various normal bioprocesses. Evidence is emerging that mA modification and its regulatory proteins also play critical roles in various cancers including leukemia, brain tumor, breast cancer and lung cancer, etc. For instance, FTO, the first mA demethylase identified, has been reported recently to play an oncogenic role in leukemia and glioblastoma. ALKBH5 (another mA demethylase) has been reported to exert a tumor-promoting function in glioblastoma and breast cancer. METTL3 (a major mA methyltransferase) likely plays distinct roles between glioblastoma and lung cancer. Here we discuss the recent progress and future prospects in study of mA machinery in cancer.
RNA mA modification and its function in diseases.
Tong Jiyu,Flavell Richard A,Li Hua-Bing
Frontiers of medicine
N-methyladenosine (mA) is the most common post-transcriptional RNA modification throughout the transcriptome, affecting fundamental aspects of RNA metabolism. mA modification could be installed by mA "writers" composed of core catalytic components (METTL3/METTL14/WTAP) and newly defined regulators and removed by mA "erasers" (FTO and ALKBH5). The function of mA is executed by mA "readers" that bind to mA directly (YTH domain-containing proteins, eIF3 and IGF2BPs) or indirectly (HNRNPA2B1). In the past few years, advances in mA modulators ("writers," "erasers," and "readers") have remarkably renewed our understanding of the function and regulation of mA in different cells under normal or disease conditions. However, the mechanism and the regulatory network of mA are still largely unknown. Moreover, investigations of the mA physiological roles in human diseases are limited. In this review, we summarize the recent advances in mA research and highlight the functional relevance and importance of mA modification in in vitro cell lines, in physiological contexts, and in cancers.
Structural basis for eukaryotic mRNA modification.
Fisher Andrew J,Beal Peter A
Current opinion in structural biology
All messenger RNAs in eukaryotes are modified co-transcriptionally and post-transcriptionally. They are all capped at the 5'-end and polyadenylated at the 3'-end. However, many mRNAs are also found to be chemically modified internally for regulation of mRNA processing, translation, stability, and to recode the message. This review will briefly summarize the structural basis for formation of the two most common modifications found at internal sites in mRNAs; methylation and deamination. The structures of the enzymes that catalyze these modifications show structural similarity to other family members within each modifying enzyme class. RNA methyltransferases, including METTL3/METTL14 responsible for N-methyladensosine (mA) formation, share a common structural core and utilize S-adenosyl methionine as a methyl donor. RNA deaminases, including adenosine deaminases acting on RNA (ADARs), also share a common structural core and similar signature sequence motif with conserved residues used for binding zinc and catalyzing the deamination reaction. In spite of recent reports of high resolution structures for members of these two RNA-modifying enzyme families, a great deal remains to be uncovered for a complete understanding of the structural basis for mRNA modification. Of particular interest is the definition of factors that control modification site specificity.
RNA N-methyladenosine modification in cancers: current status and perspectives.
Deng Xiaolan,Su Rui,Weng Hengyou,Huang Huilin,Li Zejuan,Chen Jianjun
N-methyladenosine (mA), the most abundant internal modification in eukaryotic messenger RNAs (mRNAs), has been shown to play critical roles in various normal bioprocesses such as tissue development, stem cell self-renewal and differentiation, heat shock or DNA damage response, and maternal-to-zygotic transition. The mA modification is deposited by the mA methyltransferase complex (MTC; i.e., writer) composed of METTL3, METTL14 and WTAP, and probably also VIRMA and RBM15, and can be removed by mA demethylases (i.e., erasers) such as FTO and ALKBH5. The fates of mA-modified mRNAs rely on the functions of distinct proteins that recognize them (i.e., readers), which may affect the stability, splicing, and/or translation of target mRNAs. Given the functional importance of the mA modification machinery in normal bioprocesses, it is not surprising that evidence is emerging that dysregulation of mA modification and the associated proteins also contributes to the initiation, progression, and drug response of cancers. In this review, we focus on recent advances in the study of biological functions and the underlying molecular mechanisms of dysregulated mA modification and the associated machinery in the pathogenesis and drug response of various types of cancers. In addition, we also discuss possible therapeutic interventions against the dysregulated mA machinery to treat cancers.
The Role of Dynamic m A RNA Methylation in Photobiology.
Robinson Myles,Shah Palak,Cui Yan-Hong,He Yu-Ying
Photochemistry and photobiology
N -methyladenosine (m A) is the most abundant internal RNA modification among numerous post-transcriptional modifications identified in eukaryotic mRNA. m A modification of RNA is catalyzed by the "writer" m A methyltransferase enzyme complex, consisting of METTL3, METTL14, WTAP and KIAA1429. The m A modification is reversible and can be removed by "eraser" m A demethylase enzymes, namely, FTO and ALKBH5. The biological function of m A modification on RNA is carried out by RNA-binding effector proteins called "readers." Varied functions of the reader proteins regulate mRNA metabolism by affecting stability, translation, splicing or nuclear export. The epitranscriptomic gene regulation by m A RNA methylation regulates various pathways, which contribute to basic cellular processes essential for cell maintenance, development and cell fate, and affect response to external stimuli and stressors. In this review, we summarize the recent advances in the regulation and function of m A RNA methylation, with a focus on UV-induced DNA damage response and the circadian clock machinery. Insights into the mechanisms of m A RNA regulation and post-transcriptional regulatory function in these biological processes may facilitate the development of new preventive and therapeutic strategies for various diseases related to dysregulation of UV damage response and circadian rhythm.
The Critical Role of RNA mA Methylation in Cancer.
Lan Qing,Liu Pei Y,Haase Jacob,Bell Jessica L,Hüttelmaier Stefan,Liu Tao
Since the identification of the first RNA demethylase and the establishment of methylated RNA immunoprecipitation-sequencing methodology 6 to 7 years ago, RNA methylation has emerged as a widespread phenomenon and a critical regulator of transcript expression. This new layer of regulation is termed "epitranscriptomics." The most prevalent RNA methylation, -methyladenosine (mA), occurs in approximately 25% of transcripts at the genome-wide level and is enriched around stop codons, in 5'- and 3'-untranslated regions, and within long internal exons. RNA mA modification regulates RNA splicing, translocation, stability, and translation into protein. mA is catalyzed by the RNA methyltransferases METTL3, METTL14, and METTL16 (writers), is removed by the demethylases FTO and ALKBH5 (erasers), and interacts with mA-binding proteins, such as YTHDF1 and IGF2BP1 (readers). RNA methyltransferases, demethylases, and mA-binding proteins are frequently upregulated in human cancer tissues from a variety of organ origins, increasing onco-transcript and oncoprotein expression, cancer cell proliferation, survival, tumor initiation, progression, and metastasis. Although RNA methyltransferase inhibitors are not available yet, FTO inhibitors have shown promising anticancer effects and in animal models of cancer. Further screening for selective and potent RNA methyltransferase, demethylase, or mA-binding protein inhibitors may lead to compounds suitable for future clinical trials in cancer patients.
The role of mA RNA methylation in human cancer.
Chen Xiao-Yu,Zhang Jing,Zhu Jin-Shui
N-methyladenosine (mA) is identified as the most common, abundant and conserved internal transcriptional modification, especially within eukaryotic messenger RNAs (mRNAs). MA modification is installed by the mA methyltransferases (METTL3/14, WTAP, RBM15/15B and KIAA1429, termed as "writers"), reverted by the demethylases (FTO and ALKBH5, termed as "erasers") and recognized by mA binding proteins (YTHDF1/2/3, IGF2BP1 and HNRNPA2B1, termed as "readers"). Acumulating evidence shows that, mA RNA methylation has an outsize effect on RNA production/metabolism and participates in the pathogenesis of multiple diseases including cancers. Until now, the molecular mechanisms underlying mA RNA methylation in various tumors have not been comprehensively clarified. In this review, we mainly summarize the recent advances in biological function of mA modifications in human cancer and discuss the potential therapeutic strategies.
Epigenetic Regulation of mA Modifications in Human Cancer.
Zhao Wei,Qi Xiaoqian,Liu Lina,Ma Shiqing,Liu Jingwen,Wu Jie
Molecular therapy. Nucleic acids
N6-methyladenosine (mA) is the most prevalent internal RNA modification, especially within eukaryotic messenger RNAs (mRNAs). mA modifications of RNA regulate splicing, translocation, stability, and translation into proteins. mA modifications are catalyzed by RNA methyltransferases, such as METTL3, METTL14, and WTAP (writers); the modifications are removed by the demethylases fat mass and obesity-associated protein (FTO) and ALKBH5 (ALKB homolog 5) (erasers); and the modifications are recognized by mA-binding proteins, such as YTHDF domain-containing proteins and IGF2BPs (readers). Abnormal changes in the mA levels of these genes are closely related to tumor occurrence and development. In this paper, we review the role of mA in human cancer and summarize its prospective applications in cancer.
Characterization of METTL16 as a cytoplasmic RNA binding protein.
Nance Daniel J,Satterwhite Emily R,Bhaskar Brinda,Misra Sway,Carraway Kristen R,Mansfield Kyle D
mRNA modification by N6-methyladenosine (m6A) is involved in many post-transcriptional regulation processes including mRNA stability, splicing and promotion of translation. Accordingly, the recently identified mRNA methylation complex containing METTL3, METTL14, and WTAP has been the subject of intense study. However, METTL16 (METT10D) has also been identified as an RNA m6A methyltransferase that can methylate both coding and noncoding RNAs, but its biological role remains unclear. While global studies have identified many potential RNA targets of METTL16, only a handful, including the long noncoding RNA MALAT1, the snRNA U6, as well as the mRNA MAT2A have been verified and/or studied to any great extent. In this study we identified/verified METTL16 targets by immunoprecipitation of both endogenous as well as exogenous FLAG-tagged protein. Interestingly, exogenously overexpressed METTL16 differed from the endogenous protein in its relative affinity for RNA targets which prompted us to investigate METTL16's localization within the cell. Surprisingly, biochemical fractionation revealed that a majority of METTL16 protein resides in the cytoplasm of a number of cells. Furthermore, siRNA knockdown of METTL16 resulted in expression changes of a few mRNA targets suggesting that METTL16 may play a role in regulating gene expression. Thus, while METTL16 has been reported to be a nuclear protein, our findings suggest that METTL16 is also a cytoplasmic methyltransferase that may alter its RNA binding preferences depending on its cellular localization. Future studies will seek to confirm differences between cytoplasmic and nuclear RNA targets in addition to exploring the physiological role of METTL16 through long-term knockdown.
RNA N-methyladenosine modification in solid tumors: new therapeutic frontiers.
Melstrom Laleh,Chen Jianjun
Cancer gene therapy
Epigenetic mRNA modification is an evolving field. N-methyladenosine (mA) is the most frequent internal transcriptional modification in eukaryotic messenger RNAs (mRNAs). This review will discuss the functions of the mA mRNA machinery, including its "writers" that are components of the methyltransferase complex, its "readers" and its "erasers" (specifically FTO and ALKBH5) in cancer. The writers deposit the mA and include METTL3, METTL14, WTAP, VIRMA, and RBM15. MA methylation is removed by the m6A demethylases (FTO and ALKBH5). Lastly, the most diverse members are the readers that can contribute to mRNA splicing, stability, translation, and nuclear export. Many of these functions continue to be elucidated. The dysregulation of this machinery in various malignancies and the associated impact on tumorigenesis and drug response will be discussed herein with a focus on solid tumors. It is clear that, by contributing to either mRNA stability or translation, there are downstream targets that are impacted, contributing to cancer progression and the self-renewal ability of cancer stem cells.
Role of the N6-methyladenosine RNA mark in gene regulation and its implications on development and disease.
Chandola Udita,Das Radhika,Panda Binay
Briefings in functional genomics
Epigenetics is a field that encompasses chemical modifications of DNA and histone proteins, both of which alter gene expression without changing the underlying nucleotide sequence. DNA methylation and modifications of histone tails have been studied in detail and are now known to be global gene regulatory mechanisms. An analogous post-transcriptional modification is chemical modification of specific nucleotides in RNA. Study of RNA modifications is a nascent field as yet, and the significance of these marks in controlling cell growth and differentiation is just beginning to be appreciated. The addition of a methyl group to adenosine (N-methyl-6-adenosine) or m6A is the most abundant modification in mammalian mRNAs. Though identified four decades ago, interest in this particular modification was set off by the discovery that the obesity gene FTO was an RNA demethylase. Since then, many studies have investigated m6A modification in different species. In this review, we summarize the current literature and hypotheses about the presence and function of this ubiquitous RNA modification in mammals, viruses, yeast and plants in terms of the consensus sequence and the methyltransferase/demethylation machinery identified thus far. We discuss its potential role in regulating molecular and physiological processes in each of these organisms, especially its role in RNA splicing, RNA degradation and development. We also enlist the methodologies developed so far, both locus-specific and transcriptome-wide, to study this modification. Lastly, we discuss whether m6A alterations have consequences on modulating disease aetiology, and speculate about its potential role in cancer.
The dual role of N6-methyladenosine modification of RNAs is involved in human cancers.
He Liujia,Li Jiangfeng,Wang Xiao,Ying Yufan,Xie Haiyun,Yan Huaqing,Zheng Xiangyi,Xie Liping
Journal of cellular and molecular medicine
As the most abundant and reversible RNA modification in eukaryotic cells, m A triggers a new layer of epi-transcription. M A modification occurs through a methylation process modified by "writers" complexes, reversed by "erasers", and exerts its role depending on various "readers". Emerging evidence shows that there is a strong association between m A and human diseases, especially cancers. Herein, we review bi-aspects of m A in regulating cancers mediated by the m A-associated proteins, which exert vital and specific roles in the development of various cancers. Generally, the m A modification performs promotion or inhibition functions (dual role) in tumorigenesis and progression of various cancers, which suggests a new concept in cancer regulations. In addition, m A-targeted therapies including competitive antagonists of m A-associated proteins may provide a new tumour intervention in the future.
mA mRNA modifications are deposited in nascent pre-mRNA and are not required for splicing but do specify cytoplasmic turnover.
Ke Shengdong,Pandya-Jones Amy,Saito Yuhki,Fak John J,Vågbø Cathrine Broberg,Geula Shay,Hanna Jacob H,Black Douglas L,Darnell James E,Darnell Robert B
Genes & development
Understanding the biologic role of -methyladenosine (mA) RNA modifications in mRNA requires an understanding of when and where in the life of a pre-mRNA transcript the modifications are made. We found that HeLa cell chromatin-associated nascent pre-mRNA (CA-RNA) contains many unspliced introns and mA in exons but very rarely in introns. The mA methylation is essentially completed upon the release of mRNA into the nucleoplasm. Furthermore, the content and location of each mA modification in steady-state cytoplasmic mRNA are largely indistinguishable from those in the newly synthesized CA-RNA or nucleoplasmic mRNA. This result suggests that quantitatively little methylation or demethylation occurs in cytoplasmic mRNA. In addition, only ∼10% of mAs in CA-RNA are within 50 nucleotides of 5' or 3' splice sites, and the vast majority of exons harboring mA in wild-type mouse stem cells is spliced the same in cells lacking the major mA methyltransferase Mettl3. Both HeLa and mouse embryonic stem cell mRNAs harboring mAs have shorter half-lives, and thousands of these mRNAs have increased half-lives (twofold or more) in Mettl3 knockout cells compared with wild type. In summary, mA is added to exons before or soon after exon definition in nascent pre-mRNA, and while mA is not required for most splicing, its addition in the nascent transcript is a determinant of cytoplasmic mRNA stability.
RNA mA methylation regulates the epithelial mesenchymal transition of cancer cells and translation of Snail.
Lin Xinyao,Chai Guoshi,Wu Yingmin,Li Jiexin,Chen Feng,Liu Jianzhao,Luo Guanzheng,Tauler Jordi,Du Jun,Lin Shuibin,He Chuan,Wang Hongsheng
N6-Methyladenosine (mA) modification has been implicated in the progression of several cancers. We reveal that during epithelial-mesenchymal transition (EMT), one important step for cancer cell metastasis, mA modification of mRNAs increases in cancer cells. Deletion of methyltransferase-like 3 (METTL3) down-regulates mA, impairs the migration, invasion and EMT of cancer cells both in vitro and in vivo. mA-sequencing and functional studies confirm that Snail, a key transcription factor of EMT, is involved in mA-regulated EMT. mA in Snail CDS, but not 3'UTR, triggers polysome-mediated translation of Snail mRNA in cancer cells. Loss and gain functional studies confirm that YTHDF1 mediates mA-increased translation of Snail mRNA. Moreover, the upregulation of METTL3 and YTHDF1 act as adverse prognosis factors for overall survival (OS) rate of liver cancer patients. Our study highlights the critical roles of mA on regulation of EMT in cancer cells and translation of Snail during this process.
Genome-Wide Maps of m6A circRNAs Identify Widespread and Cell-Type-Specific Methylation Patterns that Are Distinct from mRNAs.
Zhou Chan,Molinie Benoit,Daneshvar Kaveh,Pondick Joshua V,Wang Jinkai,Van Wittenberghe Nicholas,Xing Yi,Giallourakis Cosmas C,Mullen Alan C
N-methyladenosine (mA) is the most abundant internal modification of mRNAs and is implicated in all aspects of post-transcriptional RNA metabolism. However, little is known about mA modifications to circular (circ) RNAs. We developed a computational pipeline (AutoCirc) that, together with depletion of ribosomal RNA and mA immunoprecipitation, defined thousands of mA circRNAs with cell-type-specific expression. The presence of mA circRNAs is corroborated by interaction between circRNAs and YTHDF1/YTHDF2, proteins that read mA sites in mRNAs, and by reduced mA levels upon depletion of METTL3, the mA writer. Despite sharing mA readers and writers, mA circRNAs are frequently derived from exons that are not methylated in mRNAs, whereas mRNAs that are methylated on the same exons that compose mA circRNAs exhibit less stability in a process regulated by YTHDF2. These results expand our understanding of the breadth of mA modifications and uncover regulation of circRNAs through mA modification.
mA Facilitates eIF4F-Independent mRNA Translation.
Coots Ryan A,Liu Xiao-Min,Mao Yuanhui,Dong Leiming,Zhou Jun,Wan Ji,Zhang Xingqian,Qian Shu-Bing
In eukaryotic cells, protein synthesis typically begins with the binding of eIF4F to the 7-methylguanylate (mG) cap found on the 5' end of the majority of mRNAs. Surprisingly, overall translational output remains robust under eIF4F inhibition. The broad spectrum of eIF4F-resistant translatomes is incompatible with cap-independent translation mediated by internal ribosome entry sites (IRESs). Here, we report that N-methyladenosine (mA) facilitates mRNA translation that is resistant to eIF4F inactivation. Depletion of the methyltransferase METTL3 selectively inhibits translation of mRNAs bearing 5' UTR methylation, but not mRNAs with 5' terminal oligopyrimidine (TOP) elements. We identify ABCF1 as a critical mediator of mA-promoted translation under both stress and physiological conditions. Supporting the role of ABCF1 in mA-facilitated mRNA translation, ABCF1-sensitive transcripts largely overlap with METTL3-dependent mRNA targets. By illustrating the scope and mechanism of eIF4F-independent mRNA translation, these findings reshape our current perceptions of cellular translational pathways.
N6-Methyladenosine Regulates the Expression and Secretion of TGFβ1 to Affect the Epithelial-Mesenchymal Transition of Cancer Cells.
Li Jiexin,Chen Feng,Peng Yanxi,Lv Ziyan,Lin Xinyao,Chen Zhuojia,Wang Hongsheng
N6-methyladenosine (mA) is the most abundant modification on eukaryotic mRNA, which regulates all steps of the mRNA life cycle. An increasing number of studies have shown that mA methylation plays essential roles in tumor development. However, the relationship between mA and the progression of cancers remains to be explored. Here, we reported that transforming growth factor-β (TGFβ1)-induced epithelial-mesenchymal transition (EMT) was inhibited in methyltransferase-like 3 (METTL3) knockdown (Mettl3) cells. The expression of TGFβ1 was up-regulated, while self-stimulated expression of TGFβ1 was suppressed in Mettl3 cells. We further revealed that mA promoted TGFB1 mRNA decay, but impaired TGFB1 translation progress. Besides this, the autocrine of TGFβ1 was disrupted in Mettl3 cells via interrupting TGFβ1 dimer formation. Lastly, we found that Snail, which was down-regulated in Mettl3 cells, was a key factor responding to TGFβ1-induced EMT. Together, our research demonstrated that mA performed multi-functional roles in TGFβ1 expression and EMT modulation, suggesting the critical roles of mA in cancer progression regulation.
Corrigendum: Structural basis of N-adenosine methylation by the METTL3-METTL14 complex.
Wang Xiang,Feng Jing,Xue Yuan,Guan Zeyuan,Zhang Delin,Liu Zhu,Gong Zhou,Wang Qiang,Huang Jinbo,Tang Chun,Zou Tingting,Yin Ping
A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine methylation.
Liu Jianzhao,Yue Yanan,Han Dali,Wang Xiao,Fu Ye,Zhang Liang,Jia Guifang,Yu Miao,Lu Zhike,Deng Xin,Dai Qing,Chen Weizhong,He Chuan
Nature chemical biology
N(6)-methyladenosine (m(6)A) is the most prevalent and reversible internal modification in mammalian messenger and noncoding RNAs. We report here that human methyltransferase-like 14 (METTL14) catalyzes m(6)A RNA methylation. Together with METTL3, the only previously known m(6)A methyltransferase, these two proteins form a stable heterodimer core complex of METTL3-METTL14 that functions in cellular m(6)A deposition on mammalian nuclear RNAs. WTAP, a mammalian splicing factor, can interact with this complex and affect this methylation.
Discovery of Small Molecules that Activate RNA Methylation through Cooperative Binding to the METTL3-14-WTAP Complex Active Site.
Selberg Simona,Blokhina Daria,Aatonen Maria,Koivisto Pertti,Siltanen Antti,Mervaala Eero,Kankuri Esko,Karelson Mati
Chemical modifications of RNA provide an additional, epitranscriptomic, level of control over cellular functions. N-6-methylated adenosines (m6As) are found in several types of RNA, and their amounts are regulated by methyltransferases and demethylases. One of the most important enzymes catalyzing generation of m6A on mRNA is the trimer N-6-methyltransferase METTL3-14-WTAP complex. Its activity has been linked to such critical biological processes as cell differentiation, proliferation, and death. We used in silico-based discovery to identify small-molecule ligands that bind to METTL3-14-WTAP and determined experimentally their binding affinity and kinetics, as well as their effect on enzymatic function. We show that these ligands serve as activators of the METTL3-14-WTAP complex.
Structural Basis for Cooperative Function of Mettl3 and Mettl14 Methyltransferases.
Wang Ping,Doxtader Katelyn A,Nam Yunsun
N(6)-methyladenosine (m(6)A) is a prevalent, reversible chemical modification of functional RNAs and is important for central events in biology. The core m(6)A writers are Mettl3 and Mettl14, which both contain methyltransferase domains. How Mettl3 and Mettl14 cooperate to catalyze methylation of adenosines has remained elusive. We present crystal structures of the complex of Mettl3/Mettl14 methyltransferase domains in apo form as well as with bound S-adenosylmethionine (SAM) or S-adenosylhomocysteine (SAH) in the catalytic site. We determine that the heterodimeric complex of methyltransferase domains, combined with CCCH motifs, constitutes the minimally required regions for creating m(6)A modifications in vitro. We also show that Mettl3 is the catalytically active subunit, while Mettl14 plays a structural role critical for substrate recognition. Our model provides a molecular explanation for why certain mutations of Mettl3 and Mettl14 lead to impaired function of the methyltransferase complex.
METTL3 regulates WTAP protein homeostasis.
Sorci Melissa,Ianniello Zaira,Cruciani Sonia,Larivera Simone,Ginistrelli Lavinia Ceci,Capuano Ernestina,Marchioni Marcella,Fazi Francesco,Fatica Alessandro
Cell death & disease
The Wilms tumor 1 (WT1)-associated protein (WTAP) is upregulated in many tumors, including, acute myeloid leukemia (AML), where it plays an oncogenic role by interacting with different proteins involved in RNA processing and cell proliferation. In addition, WTAP is also a regulator of the nuclear complex required for the deposition of N-methyladenosine (m6A) into mRNAs, containing the METTL3 methyltransferase. However, it is not clear if WTAP may have m6A-independent regulatory functions that might contribute to its oncogenic role. Here, we show that both knockdown and overexpression of METTL3 protein results in WTAP protein upregulation, indicating that METTL3 levels are critical for WTAP protein homeostasis. However, we show that WTAP upregulation is not sufficient to promote cell proliferation in the absence of a functional METTL3. Therein, these data indicate that the reported oncogenic function of WTAP is strictly connected to a functional m6A methylation complex.
Structural basis of N(6)-adenosine methylation by the METTL3-METTL14 complex.
Wang Xiang,Feng Jing,Xue Yuan,Guan Zeyuan,Zhang Delin,Liu Zhu,Gong Zhou,Wang Qiang,Huang Jinbo,Tang Chun,Zou Tingting,Yin Ping
Chemical modifications of RNA have essential roles in a vast range of cellular processes. N(6)-methyladenosine (m(6)A) is an abundant internal modification in messenger RNA and long non-coding RNA that can be dynamically added and removed by RNA methyltransferases (MTases) and demethylases, respectively. An MTase complex comprising methyltransferase-like 3 (METTL3) and methyltransferase-like 14 (METTL14) efficiently catalyses methyl group transfer. In contrast to the well-studied DNA MTase, the exact roles of these two RNA MTases in the complex remain to be elucidated. Here we report the crystal structures of the METTL3-METTL14 heterodimer with MTase domains in the ligand-free, S-adenosyl methionine (AdoMet)-bound and S-adenosyl homocysteine (AdoHcy)-bound states, with resolutions of 1.9, 1.71 and 1.61 Å, respectively. Both METTL3 and METTL14 adopt a class I MTase fold and they interact with each other via an extensive hydrogen bonding network, generating a positively charged groove. Notably, AdoMet was observed in only the METTL3 pocket and not in METTL14. Combined with biochemical analysis, these results suggest that in the m(6)A MTase complex, METTL3 primarily functions as the catalytic core, while METTL14 serves as an RNA-binding platform, reminiscent of the target recognition domain of DNA N(6)-adenine MTase. This structural information provides an important framework for the functional investigation of m(6)A.
The m(6)A Methyltransferase METTL3 Promotes Translation in Human Cancer Cells.
Lin Shuibin,Choe Junho,Du Peng,Triboulet Robinson,Gregory Richard I
METTL3 is an RNA methyltransferase implicated in mRNA biogenesis, decay, and translation control through N(6)-methyladenosine (m(6)A) modification. Here we find that METTL3 promotes translation of certain mRNAs including epidermal growth factor receptor (EGFR) and the Hippo pathway effector TAZ in human cancer cells. In contrast to current models that invoke m(6)A reader proteins downstream of nuclear METTL3, we find METTL3 associates with ribosomes and promotes translation in the cytoplasm. METTL3 depletion inhibits translation, and both wild-type and catalytically inactive METTL3 promote translation when tethered to a reporter mRNA. Mechanistically, METTL3 enhances mRNA translation through an interaction with the translation initiation machinery. METTL3 expression is elevated in lung adenocarcinoma and using both loss- and gain-of-function studies, we find that METTL3 promotes growth, survival, and invasion of human lung cancer cells. Our results uncover an important role of METTL3 in promoting translation of oncogenes in human lung cancer.
METTL3-mediated N-methyladenosine mRNA modification enhances long-term memory consolidation.
Zhang Zeyu,Wang Meng,Xie Dongfang,Huang Zenghui,Zhang Lisha,Yang Ying,Ma Dongxue,Li Wenguang,Zhou Qi,Yang Yun-Gui,Wang Xiu-Jie
The formation of long-term memory is critical for learning ability and social behaviors of humans and animals, yet its underlying mechanisms are largely unknown. We found that the efficacy of hippocampus-dependent memory consolidation is regulated by METTL3, an RNA N-methyladenosine (mA) methyltransferase, through promoting the translation of neuronal early-response genes. Such effect is exquisitely dependent on the mA methyltransferase function of METTL3. Depleting METTL3 in mouse hippocampus reduces memory consolidation ability, yet unimpaired learning outcomes can be achieved if adequate training was given or the mA methyltransferase function of METTL3 was restored. The abundance of METTL3 in wild-type mouse hippocampus is positively correlated with learning efficacy, and overexpression of METTL3 significantly enhances long-term memory consolidation. These findings uncover a direct role of RNA mA modification in regulating long-term memory formation, and also indicate that memory efficacy difference among individuals could be compensated by repeated learning.
mRNA circularization by METTL3-eIF3h enhances translation and promotes oncogenesis.
Choe Junho,Lin Shuibin,Zhang Wencai,Liu Qi,Wang Longfei,Ramirez-Moya Julia,Du Peng,Kim Wantae,Tang Shaojun,Sliz Piotr,Santisteban Pilar,George Rani E,Richards William G,Wong Kwok-Kin,Locker Nicolas,Slack Frank J,Gregory Richard I
N-methyladenosine (mA) modification of mRNA is emerging as an important regulator of gene expression that affects different developmental and biological processes, and altered mA homeostasis is linked to cancer. mA modification is catalysed by METTL3 and enriched in the 3' untranslated region of a large subset of mRNAs at sites close to the stop codon. METTL3 can promote translation but the mechanism and relevance of this process remain unknown. Here we show that METTL3 enhances translation only when tethered to reporter mRNA at sites close to the stop codon, supporting a mechanism of mRNA looping for ribosome recycling and translational control. Electron microscopy reveals the topology of individual polyribosomes with single METTL3 foci in close proximity to 5' cap-binding proteins. We identify a direct physical and functional interaction between METTL3 and the eukaryotic translation initiation factor 3 subunit h (eIF3h). METTL3 promotes translation of a large subset of oncogenic mRNAs-including bromodomain-containing protein 4-that is also mA-modified in human primary lung tumours. The METTL3-eIF3h interaction is required for enhanced translation, formation of densely packed polyribosomes and oncogenic transformation. METTL3 depletion inhibits tumorigenicity and sensitizes lung cancer cells to BRD4 inhibition. These findings uncover a mechanism of translation control that is based on mRNA looping and identify METTL3-eIF3h as a potential therapeutic target for patients with cancer.