Multivalent m6A motifs promote phase separation of YTHDF proteins

Gao, Yifei; Pei, Gaofeng; Li, Dongxue; Li, Ru; Shao, Yanqiu; Zhang, Qiangfeng Cliff; Li, Pilong

doi:10.1038/s41422-019-0210-3

Cited by 136 publications

(114 citation statements)

References 14 publications

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“…As shown in Figure 1E, phase separation mediated by YTHDF2 increases in a manner that is dependent on the number of m6A sites in the RNA oligos (50bp RNAs containing either 0, 1, 5, or 10 m 6 A). These results are consistent with the recent reports that RNA oligos containing multiple m 6 A methylation sites robustly induce droplet formation by YTHDF2 (Gao et al,2019;Ries et al,2019). Consistently, induction of phase separation of YTHDF2 by m6A RNAs seems to be dependent on the YTH domain, as even the RNA containing 10 m 6 A failed to enhance phase separation in vitro when the m6A-binding capability of the YTH domain is compromised (YTHDF2 aa 230-579 carrying W432A, W486A) (Li et al,2014) ( Figure 1F).…”

Section: Mainsupporting

confidence: 93%

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Binding to m⁶A RNA promotes YTHDF2-mediated phase separation

Jia-hua

Wang

Diao

et al. 2019

Preprint

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As the most abundant modification on mRNA in mammal, N6-Methyladenosine (m 6 A) has been demonstrated to play important roles in various biological processes including mRNA splicing, translation and degradation. m6A reader proteins have been shown to play central roles in these processes. One of the m6A readers, YTHDF2 is localized to the P granules, which are liquid-like droplets where RNA degradation occurs. How YTHDF2 is localized to P granules is unknown. Here we provide evidence that YTHDF2 forms liquid droplets and phase separate, mediated by its low complexity (LC) domains. Interestingly, the ability to phase separate is robustly stimulated by m 6 A RNAs in vitro. In vivo, YTHDF2 phase separation may in fact be dependent on m 6 A RNA and YTHDF2 binding to m 6 A RNA, since a YTHDF2 m 6 A-binding defective mutant or a wildtype YTHDF2 assayed in cells lacking m 6 A RNAs, both fail to phase separate. The ability of phase separate is not limited to YTHDF2; we find other members of the YTH-domain m 6 A readers can also undergo phase separation. Our findings suggest that m 6 A RNA induced phase separation of m 6 A readers may play an important role in their distributions to different phase-separated compartments in cells.

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Section: Mainsupporting

confidence: 93%

“…While this manuscript was in preparation, several groups reported that YTHDF family proteins display phase separation potential (Fu and Zhuang,2019;Gao et al,2019;Ries et al,2019) stimulated by m 6 A RNA (Gao et al,2019;Ries et al,2019), consistent with our finding that m 6 A promotes the phase separation potential of YTHDF2.…”

Section: Mainsupporting

confidence: 89%

Binding to m⁶A RNA promotes YTHDF2-mediated phase separation

Jia-hua

Wang

Diao

et al. 2019

Preprint

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“…S7), suggesting that they might be guided to siRNA bodies possibly via the physical interaction with SGS3. Several other studies have shown that LLPS can be enhanced when additional biomolecules are supplemented 55,71,72 . In this regard, the functional role of the interacting proteins on the phase separation behaviour of SGS3 and the formation of siRNA bodies will be worth to be followed and investigated.…”

Section: Discussionmentioning

confidence: 96%

Translational inhibition and phase separation primes the epigenetic silencing of transposons

Kim

Lei

et al. 2020

Preprint

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19Transposons are mobile DNAs that can cause fatal mutations. To counteract these 20 genome invaders, the host genomes deploy small interfering (si) RNAs to initiate and 21 establish the epigenetic silencing. However, the regulatory mechanisms for the selective 22 recognition of transposons by the host genomes remain still elusive. Here we show that plant 23 transposon RNAs undergo frequent ribosome stalling caused by their inherently unfavourable 24 codon sequence usage. The ribosome stalling then causes the RNA truncation and the 25 localization to siRNA bodies, which are both critical prerequisites for the siRNA processing. 26 In addition, SGS3, the key protein in the siRNA biogenesis pathway, forms liquid droplets in 27 vitro through its prion-like domains implicating the role of liquid-liquid phase separation in 28 the formation of the siRNA bodies. Our study provides a novel insight into the regulatory 29 mechanisms for the recognition of invasive genetic elements which is essential for the 30 maintenance of genome integrity. 31 32 33 Keywords 34 transposon, codon optimality, ribosome stalling, easiRNA, RDR6-RdDM, liquid-liquid phase 35 separation, siRNA body, stress granule 36 37 38 45 of transposons 5 . While the canonical RdDM is mainly for reinforcing the silent state of 46 transposons, the recently proposed alternative RdDM pathway suppresses the active 47 transposons that are induced upon the epigenetic mutations and in several development 48 stages 3,6-10 . RNA-DEPENDENT RNA POLYMERASE 6 (RDR6)-SUPPRESSOR OF GENE 49 SILENCING 3 (SGS3) complex serves as a detector of transposon RNAs that selectively 50 processes them into 21 or 22-nt siRNAs (also referred to as epigenetically activated siRNAs, 51 easiRNAs) 11,12 . Transposon-derived siRNAs trigger both post-transcriptional repression by 52 cleaving the transposon RNAs (and then subsequently initiating the secondary siRNA 53 biogenesis) and establishment of the epigenetic silencing by recruiting DNA 54 methyltransferases to the target TE chromatin 5,7,8 . The biogenesis of 21 or 22-nt siRNAs in 55 plants is initiated by RDR6 which is an RNA-dependent RNA polymerase that forms double-56stranded RNAs by templating directly the target RNAs 13 . SGS3 is an RNA-binding protein 57 that interacts with RDR6 and is essential for its function 13 . The duplex RNAs are then sliced 58 to 21 or 22-nt siRNAs by DICER-LIKE 3 or 4 (DCL3/4) 8,14,15 . Since RDR6 templates are 59 directly derived from the RNAs of those mobile elements, RDR6 and the resulting easiRNAs 60 are deemed the first line of the host immune system against the invasive genetic elements. 61The RDR6-mediated easiRNA production pathway (RDR6-RdDM) is usually 62 prevented in the transcripts derived from genes by the RNA decay pathways [16][17][18] . This raises 63 4 an important question of how RDR6 specifically recognizes its TE targets and establish their 64 epigenetic silencing. Previous reports showed that the initial cleavage of target transcripts is a 65 critical prerequisite for RDR6 recog...

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“…m 6 A-containing linear and circular RNAs undergo endoribonucleolytic cleavage through YTHDF2-HRSP12-RNase P/MRP axis, coupled to CCR4-NOT complex-mediated deadenylating pathway (38) (Figure 1). In addition, a recent study reported that multivalent m 6 A-modified RNAs could promote the phase separation of YTHDFs and that phase separation of m 6 A and YTHDF2 might participate in cellular response to stresses, despite the uncertainty of its specific role (39).…”

Section: Mrna Translation or Degradation In Cytoplasmmentioning

confidence: 99%

N6-Methyladenosine: A Novel RNA Imprint in Human Cancer

Liu

et al. 2019

Front. Oncol.

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N 6 -Methyladenosine (m 6 A), a pervasive posttranscriptional modification which is reversible, has been among hotspot issues in the past several years. The balance of intracellular m 6 A levels is dynamically maintained by methyltransferase complex and demethylases. Meanwhile, m 6 A reader proteins specifically recognize modified residues and convey messages so as to set up an efficient and orderly network of m 6 A regulation. The m 6 A mark has proved to affect every step of RNA life cycle, from processing in nucleus to translation or degradation in cytoplasm. Subsequently, disorders in m 6 A methylation are directly related to aberrant RNA metabolism, which results in tumorigenesis and altered drug response. Therefore, uncovering the underlying mechanism of m 6 A in oncogenic transformation and tumor progression seeks opportunities for novel targets in cancer therapy. In this review, we conclude the extensive impact of m 6 A on RNA metabolism and highlight its relevance with human cancer, implicating the far-reaching value in clinical application.

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Multivalent m6A motifs promote phase separation of YTHDF proteins

Cited by 136 publications

References 14 publications

Binding to m⁶A RNA promotes YTHDF2-mediated phase separation

Binding to m⁶A RNA promotes YTHDF2-mediated phase separation

Translational inhibition and phase separation primes the epigenetic silencing of transposons

N6-Methyladenosine: A Novel RNA Imprint in Human Cancer

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Multivalent m6A motifs promote phase separation of YTHDF proteins

Cited by 136 publications

References 14 publications

Binding to m6A RNA promotes YTHDF2-mediated phase separation

Binding to m6A RNA promotes YTHDF2-mediated phase separation

Translational inhibition and phase separation primes the epigenetic silencing of transposons

N6-Methyladenosine: A Novel RNA Imprint in Human Cancer

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Binding to m⁶A RNA promotes YTHDF2-mediated phase separation

Binding to m⁶A RNA promotes YTHDF2-mediated phase separation