YTHDF2 mediates the mRNA degradation of the tumor suppressors to induce AKT phosphorylation in N6-methyladenosine-dependent way in prostate cancer.
YTHDF2 mediates the mRNA degradation of the tumor suppressors to induce AKT phosphorylation in N6-methyladenosine-dependent way in prostate cancer.
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YTHDF2 在前列腺癌中介导肿瘤抑制因子的 mRNA 降解，以 N6-甲基腺苷依赖性方式诱导 AKT 磷酸化
DOI：
10.1186/s12943-020-01267-6
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发表时间：
2020-10-29
期刊：
影响因子：
37.3
通讯作者：
Xie L
中科院分区：
文献类型：
--
作者：
Li J;Xie H;Ying Y;Chen H;Yan H;He L;Xu M;Xu X;Liang Z;Liu B;Wang X;Zheng X;Xie L
N6 - 甲基腺苷（m6A）是人类mRNA中最丰富的修饰。新出现的证据支持m6A广泛参与各种疾病，尤其是癌症这一事实。作为关键的“读取器”，YTHDF2通常以m6A依赖的方式介导m6A修饰的mRNA的降解。然而，m6A尤其是YTHDF2在前列腺癌（PCa）中的功能和机制仍然不清楚。
为了研究YTHDF2在PCa中的功能和机制，进行了体外、体内生物功能测定和表观遗传学实验。利用基于慢病毒的shRNA技术建立了YTHDF2和METTL3的内源性表达沉默。进行集落形成、流式细胞术和Trans - well实验以鉴定细胞功能。将皮下异种移植和转移小鼠模型与体内成像系统相结合，以研究敲低YTHDF2和METTL3时的表型。m6A RNA免疫沉淀（MeRIP）测序、mRNA测序、RIP - RT - qPCR和生物信息学分析主要用于筛选和验证YTHDF2和METTL3的直接共同靶点。此外，还使用TCGA数据库分析了YTHDF2、METTL3以及共同靶点LHPP在PCa中的表达模式及其与临床预后的相关性。
PCa中上调的YTHDF2和METTL3预示着更差的总生存率。敲低YTHDF2或METTL3在体内和体外显著抑制PCa的增殖和迁移。LHPP和NKX3 - 1被确定为YTHDF2和METTL3的直接靶点。YTHDF2直接结合到LHPP和NKX3 - 1的m6A修饰位点以介导mRNA降解。敲低YTHDF2或METTL3在mRNA和蛋白质水平上显著诱导LHPP和NKX3 - 1的表达，同时抑制磷酸化AKT。LHPP和NKX3 - 1的过表达呈现出与敲低YTHDF2或METTL3一致的表型以及AKT磷酸化抑制。因此，磷酸化AKT被确认为METTL3/YTHDF2/LHPP/NKX3 - 1的下游，可诱导肿瘤增殖和迁移。
我们提出了一种新的调控机制，即YTHDF2以m6A依赖的方式介导肿瘤抑制因子LHPP和NKX3 - 1的mRNA降解，从而调节AKT磷酸化诱导的前列腺癌肿瘤进展。我们希望我们的发现能为前列腺癌生物学提供新的概念。
BackgroundN6-methyladenosine (m6A) is the most abundant modification in mRNA of humans. Emerging evidence has supported the fact that m6A is comprehensively involved in various diseases especially cancers. As a crucial reader, YTHDF2 usually mediates the degradation of m6A-modified mRNAs in m6A-dependent way. However, the function and mechanisms of m6A especially YTHDF2 in prostate cancer (PCa) still remain elusive.MethodsTo investigate the functions and mechanisms of YTHDF2 in PCa, in vitro,in vivo biofunctional assays and epigenetics experiments were performed. Endogenous expression silencing of YTHDF2 and METTL3 was established with lentivirus-based shRNA technique. Colony formation, flow cytometry and trans-well assays were performed for cell function identifications. Subcutaneous xenografts and metastatic mice models were combined with in vivo imaging system to investigate the phenotypes when knocking down YTHDF2 and METTL3. m6A RNA immunoprecipitation (MeRIP) sequencing, mRNA sequencing, RIP-RT-qPCR and bioinformatics analysis were mainly used to screen and validate the direct common targets of YTHDF2 and METTL3. In addition, TCGA database was also used to analyze the expression pattern of YTHDF2, METTL3 and the common target LHPP in PCa, and their correlation with clinical prognosis.ResultsThe upregulated YTHDF2 and METTL3 in PCa predicted a worse overall survival rate. Knocking down YTHDF2 or METTL3 markedly inhibited the proliferation and migration of PCa in vivo and in vitro. LHPP and NKX3–1 were identified as the direct targets of both YTHDF2 and METTL3. YTHDF2 directly bound to the m6A modification sites of LHPP and NKX3–1 to mediate the mRNA degradation. Knock-down of YTHDF2 or METTL3 significantly induced the expression of LHPP and NKX3–1 at both mRNA and protein level with inhibited phosphorylated AKT. Overexpression of LHPP and NKX3–1 presented the consistent phenotypes and AKT phosphorylation inhibition with knock-down of YTHDF2 or METTL3. Phosphorylated AKT was consequently confirmed as the downstream of METTL3/YTHDF2/LHPP/NKX3–1 to induce tumor proliferation and migration.ConclusionWe propose a novel regulatory mechanism in which YTHDF2 mediates the mRNA degradation of the tumor suppressors LHPP and NKX3–1 in m6A-dependent way to regulate AKT phosphorylation-induced tumor progression in prostate cancer. We hope our findings may provide new concepts of PCa biology.
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影响因子：
16.6
作者：
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通讯作者：
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影响因子：
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作者：
Li J;Meng S;Xu M;Wang S;He L;Xu X;Wang X;Xie L
通讯作者：
Xie L
DOI：
10.1073/pnas.1721371115
发表时间：
2018-06-05
影响因子：
11.1
作者：
Fustin JM;Kojima R;Itoh K;Chang HY;Ye S;Zhuang B;Oji A;Gibo S;Narasimamurthy R;Virshup D;Kurosawa G;Doi M;Manabe I;Ishihama Y;Ikawa M;Okamura H
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影响因子：
3.7
作者：
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影响因子：
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作者：
Lei, Qunying;Jiao, Jing;Wu, Hong
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Wu, Hong