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N6-Adenosine methylation on mRNA is recognized by YTH2 domain protein of human malaria parasite Plasmodium falciparum.
Epigenetics & Chromatin ( IF 3.9 ) Pub Date : 2020-08-31 , DOI: 10.1186/s13072-020-00355-7
Gayathri Govindaraju 1, 2 , Rajashekar Varma Kadumuri 3 , Devadathan Valiyamangalath Sethumadhavan 1, 2 , C A Jabeena 1, 2 , Sreenivas Chavali 3 , Arumugam Rajavelu 1
Affiliation  

Plasmodium falciparum exhibits high translational plasticity during its development in RBCs, yet the regulation at the post-transcriptional level is not well understood. The N6-methyl adenosine (m6A) is an important epigenetic modification primarily present on mRNA that controls the levels of transcripts and efficiency of translation in eukaryotes. Recently, the dynamics of m6A on mRNAs at all three developmental stages of P. falciparum in RBCs have been profiled; however, the proteins that regulate the m6A containing mRNAs in the parasites are unknown. Using sequence analysis, we computationally identified that the P. falciparum genome encodes two putative YTH (YT521-B Homology) domain-containing proteins, which could potentially bind to m6A containing mRNA. We developed a modified methylated RNA immunoprecipitation (MeRIP) assay using PfYTH2 and find that it binds selectively to m6A containing transcripts. The PfYTH2 has a conserved aromatic amino acid cage that forms the methyl-binding pocket. Through site-directed mutagenesis experiments and molecular dynamics simulations, we show that F98 residue is important for m6A binding on mRNA. Fluorescence depolarization assay confirmed that PfYTH2 binds to methylated RNA oligos with high affinity. Further, MeRIP sequencing data revealed that PfYTH2 has more permissive sequence specificity on target m6A containing mRNA than other known eukaryotic YTH proteins. Taken together, here we identify and characterize PfYTH2 as the major protein that could regulate m6A containing transcripts in P. falciparum. Plasmodium spp. lost the canonical m6A-specific demethylases in their genomes, however, the YTH domain-containing proteins seem to be retained. This study presents a possibility that the YTH proteins are involved in post-transcriptional control in P. falciparum, and might orchestrate the translation of mRNA in various developmental stages of P. falciparum. This is perhaps the first characterization of the methyl-reading function of YTH protein in any parasites.

中文翻译:

mRNA 上的 N6-腺苷甲基化被人类疟原虫恶性疟原虫的 YTH2 域蛋白识别。

恶性疟原虫在红细胞发育过程中表现出高度的翻译可塑性,但转录后水平的调节尚不清楚。N6-甲基腺苷 (m6A) 是一种重要的表观遗传修饰,主要存在于 mRNA 上,可控制真核生物中的转录水平和翻译效率。最近,已经对红细胞中恶性疟原虫所有三个发育阶段的 mRNA 上 m6A 的动态进行了分析。然而,在寄生虫中调节含有 m6A 的 mRNA 的蛋白质是未知的。使用序列分析,我们通过计算确定恶性疟原虫基因组编码两个推定的包含 YTH(YT521-B 同源)结构域的蛋白质,它们可能与含有 m6A 的 mRNA 结合。我们使用 PfYTH2 开发了一种改良的甲基化 RNA 免疫沉淀 (MeRIP) 测定,发现它选择性地结合含有 m6A 的转录本。PfYTH2 有一个保守的芳香族氨基酸笼,形成甲基结合口袋。通过定点诱变实验和分子动力学模拟,我们表明 F98 残基对于 m6A 与 mRNA 的结合很重要。荧光去极化试验证实 PfYTH2 以高亲和力与甲基化 RNA 寡核苷酸结合。此外,MeRIP 测序数据显示,与其他已知的真核 YTH 蛋白相比,PfYTH2 对含有 mRNA 的靶标 m6A 具有更高的许可序列特异性。总之,在这里我们确定并表征 PfYTH2 作为可以调节恶性疟原虫中含有 m6A 的转录物的主要蛋白质。疟原虫 失去了基因组中典型的 m6A 特异性去甲基化酶,然而,含有 YTH 结构域的蛋白质似乎被保留了下来。该研究表明,YTH 蛋白可能参与恶性疟原虫的转录后调控,并可能在恶性疟原虫的各个发育阶段协调 mRNA 的翻译。这可能是对任何寄生虫中 YTH 蛋白的甲基读取功能的首次表征。
更新日期:2020-08-31
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