Dynamic assembly of the mRNA m6A methyltransferase complex is regulated by METTL3 phase separation.
Dynamic assembly of the mRNA m6A methyltransferase complex is regulated by METTL3 phase separation.
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DOI：
10.1371/journal.pbio.3001535
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发表时间：
2022-03
期刊：
影响因子：
9.8
通讯作者：
Kosik KS
中科院分区：
文献类型：
--
作者：
Han D;Longhini AP;Zhang X;Hoang V;Wilson MZ;Kosik KS
m6 A甲基化是mRNA上最丰富和可逆的化学修饰，约四分之一的真核mRNA含有至少一个m6 A修饰的碱基。mRNA m6 A甲基转移酶作家复合物相分离的核斑点的招聘可能是至关重要的，在其监管，但是，控制复杂的活动仍然不清楚。通过我们的观察，即甲基转移酶复合物的核心催化亚基，胃L3，是内源性共定位在核斑点以及在noncolocalized斑点，我们跟踪了复合物的组分与Cry 2-胃L3融合构建解开关键结构域和胃L3的相分离所需的相互作用。胃L3能够自我相互作用，并可能提供多价性以驱动缩合。细胞中的冷凝物必然包含无数的组分，每种组分都具有分配系数，这些分配系数建立了一个熵屏障，可以调节进入冷凝物的入口。在这方面，我们发现，与在扩散相和密相中的胃L14与胃L3的组成性结合相反，WTAP仅与密相中的胃L3相互作用，从而将稀相中的胃L3/胃L14单一复合物与胃L3/胃L14多组分缩合物区分开来。最后，通过其小分子辅因子S-腺苷甲硫氨酸（SAM）来控制胃L3/胃L14缩合，SAM调节两个门环的构象，并且门环附近的一些癌症相关突变可以损害胃L3缩合。因此，SAM绑定和控制作家复杂的相态之间的联系表明，其相态的调节是其功能调节的一个潜在的关键方面。大约四分之一的真核生物mRNA至少含有一个m6 A修饰的碱基，但这是如何调控的？这项研究表明，细胞可以使用液-液相分离来调节mRNA m6 A甲基转移酶复合物（胃L3/胃L14/WTAP）的动态组装，其化学计量以底物结合依赖性方式依赖于冷凝物分配。
m6A methylation is the most abundant and reversible chemical modification on mRNA with approximately one-fourth of eukaryotic mRNAs harboring at least one m6A-modified base. The recruitment of the mRNA m6A methyltransferase writer complex to phase-separated nuclear speckles is likely to be crucial in its regulation; however, control over the activity of the complex remains unclear. Supported by our observation that a core catalytic subunit of the methyltransferase complex, METTL3, is endogenously colocalized within nuclear speckles as well as in noncolocalized puncta, we tracked the components of the complex with a Cry2-METTL3 fusion construct to disentangle key domains and interactions necessary for the phase separation of METTL3. METTL3 is capable of self-interaction and likely provides the multivalency to drive condensation. Condensates in cells necessarily contain myriad components, each with partition coefficients that establish an entropic barrier that can regulate entry into the condensate. In this regard, we found that, in contrast to the constitutive binding of METTL14 to METTL3 in both the diffuse and the dense phase, WTAP only interacts with METTL3 in dense phase and thereby distinguishes METTL3/METTL14 single complexes in the dilute phase from METTL3/METTL14 multicomponent condensates. Finally, control over METTL3/METTL14 condensation is determined by its small molecule cofactor, S-adenosylmethionine (SAM), which regulates conformations of two gate loops, and some cancer-associated mutations near gate loops can impair METTL3 condensation. Therefore, the link between SAM binding and the control of writer complex phase state suggests that the regulation of its phase state is a potentially critical facet of its functional regulation. Approximately one-fourth of eukaryotic mRNAs harbor at least one m6A-modified base, but how is this regulated? This study shows that cells can use liquid-liquid phase separation to regulate dynamic assembly of the mRNA m6A methyltransferase complex (METTL3/METTL14/WTAP), with stoichiometries that depend on condensate partitioning in a substrate binding-dependent manner.
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影响因子：
2.9
作者：
Akrap N;Seidel T;Barisas BG
通讯作者：
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