目的:探讨甲基化转移酶3(methyltransferase-like 3,METTL3)联合程序死亡配体1(programmed death ligand 1,PD-L1检测在非小细胞肺癌(non-small cell lung cancer,NSCLC)患者程序性细胞死亡蛋白1(programmed death cell protein 1,PD-1...目的:探讨甲基化转移酶3(methyltransferase-like 3,METTL3)联合程序死亡配体1(programmed death ligand 1,PD-L1检测在非小细胞肺癌(non-small cell lung cancer,NSCLC)患者程序性细胞死亡蛋白1(programmed death cell protein 1,PD-1)抑制剂治疗效果和预后预测中的价值。方法:收集NSCLC患者原发病灶组织蜡块标本,免疫组织化学检测PD-L1、METTL3的蛋白表达情况,并分析其与NSCLC患者PD-1抑制剂治疗效果和预后间的联系。结果:符合入组标准的患者共228例;其中,METTL3蛋白的表达情况与病理类型、美国东部肿瘤合作组织(Eastern American Oncology Collaborative Group,ECOG)评分、治疗方案、治疗效果以及实体瘤疗效评价标准(response evaluation criteria in solid tumors,RECIST)等因素有关;METTL3蛋白高表达患者的中位无进展生存期(progression-free-survival,PFS)为12.33个月,METTL3蛋白低或无表达患者的中位PFS为6.50个月;在接受PD-1抑制剂治疗的NSCLC患者中,METTL3蛋白高表达患者有较长的PFS;METTL3蛋白高表达患者的中位总体生存期(overall-survival,OS)为23.54个月,METTL3蛋白低或无表达患者的中位OS为20.93个月,METTL3蛋白高表达与较长的OS相关。结论:METTL3蛋白可能为预测NSCLC PD-1抑制剂免疫治疗效果的标志物。展开更多
目的探讨METTL14通过调控巨噬细胞分化抑制宫颈癌病理性发展及相关机制。方法检测宫颈癌病变样本METTL14 m RNA和蛋白,以及IL-6、iNOS、Arg-1和CD206表达变化。PMA诱导THP-1细胞转化为巨噬细胞,慢病毒过表达或抑制METTL14表达,检测IL-6...目的探讨METTL14通过调控巨噬细胞分化抑制宫颈癌病理性发展及相关机制。方法检测宫颈癌病变样本METTL14 m RNA和蛋白,以及IL-6、iNOS、Arg-1和CD206表达变化。PMA诱导THP-1细胞转化为巨噬细胞,慢病毒过表达或抑制METTL14表达,检测IL-6、iNO、Arg-1和CD206表达变化以及PI3K/AKT/GSK3β/β-catenin信号通路相关蛋白表达情况。随后加入PI3K/AKT/GSK3β/β-catenin信号通路激动剂和抑制剂,检测过表达或抑制METTL14后,巨噬细胞IL-6、iNO、Arg-1和CD206表达变化,并取其上清制成条件培养基,孵育Hela细胞,检测细胞凋亡和增殖情况。结果1)宫颈癌病变组织中METTL14 mRNA和蛋白表达降低(P<0.05),巨噬细胞M1型标志物IL-6和iNOS表达明显降低(P<0.05),而M2型标志物Arg-1和CD206表达明显升高(P<0.05)。2)巨噬细胞过表达METTL14后,IL-6和iNOS表达明显升高(P<0.05),而Arg-1和CD206表达明显降低(P<0.05),M1/M2比例升高;抑制METTL14表达后,M1型标志物降低(P<0.05),M2型标志物升高(P<0.05),M1/M2比例降低。3)巨噬细胞中转染OE-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被抑制(P<0.05);加入PI3K/AKT激动剂后,M1型标志物降低而M2型标记物升高(P<0.05),M1/M2比例降低;OE-METTL14可逆转此趋势。Sh-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被激活(P<0.05),加入PI3K/AKT抑制剂后,M1型标志物升高而M2型标记物降低(P<0.05),M1/M2比例升高;Sh-METTL14可逆转此趋势。4)取转染OE-METTL14慢病毒后的巨噬细胞上清培养Hela细胞,可见细胞凋亡明显增多(P<0.05),增殖明显减少(P<0.05)。Sh-METTL14组的Hela细胞则表现出细胞凋亡减少(P<0.05),增殖增多(P<0.05)。结论METTL14通过PI3K/AKT/GSK3β/β-catenin信号通路调控巨噬细胞分化可能有促进宫颈癌细胞凋亡,抑制增殖的作用。展开更多
In non-small cell lung cancer(NSCLC),the heterogeneity promotes drug resistance,and the restricted expression of programmed death-ligand 1(PD-L1)limits the immunotherapy benefits.Based on the mechanisms related to tra...In non-small cell lung cancer(NSCLC),the heterogeneity promotes drug resistance,and the restricted expression of programmed death-ligand 1(PD-L1)limits the immunotherapy benefits.Based on the mechanisms related to translation regulation and the association with PD-L1 of methyltransferaselike 3(METTL3),the novel small-molecule inhibitor STM2457 is assumed to be useful for the treatment of NSCLC.We evaluated the efficacy of STM2457 in vivo and in vitro and confirmed the effects of its inhibition on disease progression.Next,we explored the effect of STM2457 on METTL3 and revealed its effects on the inhibition of catalytic activity and upregulation of METTL3 protein expression.Importantly,we described the genome-wide characteristics of multiple omics data acquired from RNA sequencing,ribosome profiling,and methylated RNA immunoprecipitation sequencing data under STM2457 treatment or METTL3 knockout.We also constructed a model for the regulation of the translation of METTL3 and PD-L1.Finally,we found PD-L1 upregulation by STM2457 in vivo and in vitro.In conclusion,STM2457 is a potential novel suppressor based on its inhibitory effect on tumor progression and may be able to overcome the heterogeneity based on its impact on the translatome.Furthermore,it can improve the immunotherapy outcomes based on PDL1 upregulation in NSCLC.展开更多
N6-methyladenosine(m^(6)A)plays crucial roles in development and cellular reprogramming.During embryonic development,pluripotency transitions from a naïve to a primed state,and modeling the reverse primed-to-na&#...N6-methyladenosine(m^(6)A)plays crucial roles in development and cellular reprogramming.During embryonic development,pluripotency transitions from a naïve to a primed state,and modeling the reverse primed-to-naïve transition(PNT)provides a valuable framework for investigating pluripotency regulation.Here,we show that inhibiting METTL3 significantly promotes PNT in an m^(6)A-dependent manner.Mechanistically,we found that suppressing METTL3 and YTHDF2 prolongs the lifetimes of pluripotency-associated mRNAs,such as Nanog and Sox2,during PNT.In addition,Gstp1 was identified as a downstream target of METTL3 inhibition and YTHDF2 knockout.Gstp1 overexpression enhances PNT,whereas its inhibition impedes the transition.Overall,our findings suggest that YTHDF2 facilitates the removal of pluripotency gene transcripts and Gstp1,thereby promoting PNT reprogramming through m^(6)A-mediated posttranscriptional control.展开更多
The development and homeostasis of intestinal epithelium are mediated by actively proliferating Lgr5^(+)stem cells,which pos-sess a remarkable self-renewal and differentiation capacity.Recently,our study demonstrated ...The development and homeostasis of intestinal epithelium are mediated by actively proliferating Lgr5^(+)stem cells,which pos-sess a remarkable self-renewal and differentiation capacity.Recently,our study demonstrated that N^(6)-methyladenosine(m^(6)A)methylation was essential for the survival of colonic stem cells.Here,we show that methyltransferase-like 3(METTL3)expression is downregulated in the colon mucosa in ulcerative colitis(UC)patients and strongly associated with the differentiation and maturation of goblet cells during inflammation.In mice,depletion of Mettl3 significantly inhibits the self-renewal and differentiation of Lgr5^(+)stem cells,especially the differentiation and maturation of goblet cells,resulting in intestinal dysplasia and spontaneous inflammation.Mechanistically,Mettl3 deletion-mediated m^(6)A loss facilitates the expression levels of growth factor receptor binding protein 10(Grb10)and interferon-related developmental regulator 1(Ifrd1)via increasing their messenger RNA stability.We further demonstrate that the levels of GRB10 and IFRD1 are negatively correlated with METTL3 level in UC samples.Collectively,our data indicate that METTL3 enhances the self-renewal and differentiation of Lgr5^(+)stem cells during intestinal development and inflammation,and thus it may be a potential therapeutic target for UC treatment.展开更多
Intestinal epithelial cells(IECs)are pivotal for maintaining intestinal homeostasis through self-renewal,proliferation,differentiation,and regulated cell death.While apoptosis and necroptosis are recognized as distinc...Intestinal epithelial cells(IECs)are pivotal for maintaining intestinal homeostasis through self-renewal,proliferation,differentiation,and regulated cell death.While apoptosis and necroptosis are recognized as distinct pathways,their intricate interplay remains elusive.In this study,we report that Mettl3-mediatedm6A modification maintains intesti-nal homeostasis by impeding epithelial cell death.Mettl3 knockout induces both apoptosis and necroptosis in IECs.Targeting different modes of cell death with specific inhibitors unveils that RIPK1 kinase activity is critical for the cell death triggered by Mettl3 knockout.Mechanistically,this occurs via them6A-mediated transcriptional regulation of Atf3,a transcription factor that directly binds to Cflar,the gene encoding the anti-cell death protein cFLIP.cFLIP inhibits RIPK1 activity,thereby suppressing downstream apoptotic and necroptotic signaling.Together,these find-ings delineate the essential role of the METTL3-ATF3-cFLIP axis in homeostatic regulation of the intestinal epithelium by blocking RIPK1 activity.展开更多
Background:Cancer cells selectively promote the translation of oncogenic tran-scripts to stimulate cancer progression.Although growing evidence has revealed that tRNA modifications and related genes participate in thi...Background:Cancer cells selectively promote the translation of oncogenic tran-scripts to stimulate cancer progression.Although growing evidence has revealed that tRNA modifications and related genes participate in this process,their roles in head and neck squamous cell carcinoma(HNSCC)remain largely unchar-acterized.Here,we sought to investigate the function and mechanisms of the transfer RNA(tRNA)N7-methylguanosine(m'G)modification in regulating the occurrence and development of HNSCC.Methods:Cell lost of-function and gain-of function assays,xenograft models,conditional knockout and knockin mouse models were used to study the physi-ological functions of tRNA m'G modification in HNSCC tumorigenesis.tRNA modification and expression profiling,mRNA translation profiling and res-cue assays were performed to uncover the underlying molecular mechanisms.Single-cell RNA sequencing(scRNA seq)was conducted to explore the tumor microenvironment changes.Results:The tRNA.m7G methyltransferase complex components Methyltransferase-like 1(METTL1)/WD repeat domain 4(WDR4)were upregulated in HNSCC and associated with a poor prognosis.Functionally,METTL1/WDR4 promoted HNSCC progression and metastasis in cell-based and transgenic mouse models.Mechanistically,ablation of METTL1 reduced the m'G levels of 16 tRNAS,inhibiting the translation of a subset of oncogenic transcripts,including genes related to the phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/AKT/mTOR)signaling pathway.In addition,chemical modulators of the PI3K/Akt/mTOR signaling pathway reversed the effects of Mettll in mouse HNSCC.Furthermore,scRNA-seq results revealed that Mettll knockout in mouse tumor cells altered the immune landscape and cell-cell interaction between the tumor and stromal compartment.Conclusions:The tRNA m?G methyltransferase METTLI was found to promote the development and malignancy of HNSCC through regulating global mRNA translation,including the PI3K/AKT/mTOR signaling pathway,and found to alter immune landscape.METTLI could be a promising treatment target for HNSCC patients.展开更多
Background:Methyltransferase 3(METTL3)-mediated N6-methyladenosine(m^(6)A)RNA modification has been demonstrated to be a potential factor in promoting gastric cancer(GC).METTL3 regulates a series of signaling pathways...Background:Methyltransferase 3(METTL3)-mediated N6-methyladenosine(m^(6)A)RNA modification has been demonstrated to be a potential factor in promoting gastric cancer(GC).METTL3 regulates a series of signaling pathways by modifying various mRNAs.This study aimed to identify novel METTL3-mediated signaling pathways and explored possible targets for use in the clinical setting of gastric cancer.Methods:To investigate the proliferation and metastatic capacity ofGCcell lines with METTL3 knockdown,a xenograft,lung metastasis,and popliteal lymph node metastasis model was used.Them^(6)A-modified RNA immunoprecipitation(Me-RIP)sequence was utilized to explore the target mRNAs of METTL3.Cell counting kit 8 and transwell assays were performed to investigate the promoting function of pre-B cell leukemia homeobox 1(PBX1)and GTP cyclohydrolase 1(GCH1).Western blotting and chromatin immunoprecipitation were employed to confirm the involvement of the METTL3-PBX1-GCH1 axis.ELISA and liquid chromatography-mass spectrometry were used to explore the biological function of tetrahydrobiopterin(BH_(4)).Results:Knockdown of METTL3 suppressed xenograft tumor growth and lung/lymph node metastasis in vivo.Mechanistically,we found that METTL3 combined with and stabilized PBX1 mRNAs.Chromatin immunoprecipitation(ChIP)and further experiments suggested that PBX1 acted as a transcription factor inducing GCH1 expression.Moreover,the METTL3-PBX1-GCH1 axis increased BH_(4)levels in GC cells,thereby promoting tumor progression.Conclusions:This study suggested that METTL3 enzymes promote tumor growth and lung/lymph node metastasis via METTL3-PBX1-GCH1 axis increasing BH_(4)levels in GC.展开更多
N6-methyladenosine(m^(6)A)is one of the most abundant modifications on m RNAs and plays important roles in various biological processes.The formation of m^(6)A is catalyzed by a methyltransferase complex(MTC)containin...N6-methyladenosine(m^(6)A)is one of the most abundant modifications on m RNAs and plays important roles in various biological processes.The formation of m^(6)A is catalyzed by a methyltransferase complex(MTC)containing a key factor methyltransferase-like 3(Mettl3).However,the functions of Mettl3 and m^(6)A modification in hepatic lipid and glucose metabolism remain unclear.Here,we showed that both Mettl3 expression and m^(6)A level increased in the livers of mice with high fat diet(HFD)-induced metabolic disorders.Overexpression of Mettl3 aggravated HFDinduced liver metabolic disorders and insulin resistance.In contrast,hepatocyte-specific knockout of Mettl3 significantly alleviated HFD-induced metabolic disorders by slowing weight gain,reducing lipid accumulation,and improving insulin sensitivity.Mechanistically,Mettl3 depletion-mediated m^(6)A loss caused extended RNA half-lives of metabolism-related genes,which consequently protected mice against HFD-induced metabolic syndrome.Our findings reveal a critical role of Mettl3-mediated m^(6)A in HFD-induced metabolic disorders and hepatogenous diabetes.展开更多
文摘目的:探讨甲基化转移酶3(methyltransferase-like 3,METTL3)联合程序死亡配体1(programmed death ligand 1,PD-L1检测在非小细胞肺癌(non-small cell lung cancer,NSCLC)患者程序性细胞死亡蛋白1(programmed death cell protein 1,PD-1)抑制剂治疗效果和预后预测中的价值。方法:收集NSCLC患者原发病灶组织蜡块标本,免疫组织化学检测PD-L1、METTL3的蛋白表达情况,并分析其与NSCLC患者PD-1抑制剂治疗效果和预后间的联系。结果:符合入组标准的患者共228例;其中,METTL3蛋白的表达情况与病理类型、美国东部肿瘤合作组织(Eastern American Oncology Collaborative Group,ECOG)评分、治疗方案、治疗效果以及实体瘤疗效评价标准(response evaluation criteria in solid tumors,RECIST)等因素有关;METTL3蛋白高表达患者的中位无进展生存期(progression-free-survival,PFS)为12.33个月,METTL3蛋白低或无表达患者的中位PFS为6.50个月;在接受PD-1抑制剂治疗的NSCLC患者中,METTL3蛋白高表达患者有较长的PFS;METTL3蛋白高表达患者的中位总体生存期(overall-survival,OS)为23.54个月,METTL3蛋白低或无表达患者的中位OS为20.93个月,METTL3蛋白高表达与较长的OS相关。结论:METTL3蛋白可能为预测NSCLC PD-1抑制剂免疫治疗效果的标志物。
文摘目的探讨METTL14通过调控巨噬细胞分化抑制宫颈癌病理性发展及相关机制。方法检测宫颈癌病变样本METTL14 m RNA和蛋白,以及IL-6、iNOS、Arg-1和CD206表达变化。PMA诱导THP-1细胞转化为巨噬细胞,慢病毒过表达或抑制METTL14表达,检测IL-6、iNO、Arg-1和CD206表达变化以及PI3K/AKT/GSK3β/β-catenin信号通路相关蛋白表达情况。随后加入PI3K/AKT/GSK3β/β-catenin信号通路激动剂和抑制剂,检测过表达或抑制METTL14后,巨噬细胞IL-6、iNO、Arg-1和CD206表达变化,并取其上清制成条件培养基,孵育Hela细胞,检测细胞凋亡和增殖情况。结果1)宫颈癌病变组织中METTL14 mRNA和蛋白表达降低(P<0.05),巨噬细胞M1型标志物IL-6和iNOS表达明显降低(P<0.05),而M2型标志物Arg-1和CD206表达明显升高(P<0.05)。2)巨噬细胞过表达METTL14后,IL-6和iNOS表达明显升高(P<0.05),而Arg-1和CD206表达明显降低(P<0.05),M1/M2比例升高;抑制METTL14表达后,M1型标志物降低(P<0.05),M2型标志物升高(P<0.05),M1/M2比例降低。3)巨噬细胞中转染OE-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被抑制(P<0.05);加入PI3K/AKT激动剂后,M1型标志物降低而M2型标记物升高(P<0.05),M1/M2比例降低;OE-METTL14可逆转此趋势。Sh-METTL14慢病毒组PI3K/AKT/GSK3β/β-catenin信号通路被激活(P<0.05),加入PI3K/AKT抑制剂后,M1型标志物升高而M2型标记物降低(P<0.05),M1/M2比例升高;Sh-METTL14可逆转此趋势。4)取转染OE-METTL14慢病毒后的巨噬细胞上清培养Hela细胞,可见细胞凋亡明显增多(P<0.05),增殖明显减少(P<0.05)。Sh-METTL14组的Hela细胞则表现出细胞凋亡减少(P<0.05),增殖增多(P<0.05)。结论METTL14通过PI3K/AKT/GSK3β/β-catenin信号通路调控巨噬细胞分化可能有促进宫颈癌细胞凋亡,抑制增殖的作用。
基金supported by the National Natural Science Foundation of China(Grant No.:82072593).
文摘In non-small cell lung cancer(NSCLC),the heterogeneity promotes drug resistance,and the restricted expression of programmed death-ligand 1(PD-L1)limits the immunotherapy benefits.Based on the mechanisms related to translation regulation and the association with PD-L1 of methyltransferaselike 3(METTL3),the novel small-molecule inhibitor STM2457 is assumed to be useful for the treatment of NSCLC.We evaluated the efficacy of STM2457 in vivo and in vitro and confirmed the effects of its inhibition on disease progression.Next,we explored the effect of STM2457 on METTL3 and revealed its effects on the inhibition of catalytic activity and upregulation of METTL3 protein expression.Importantly,we described the genome-wide characteristics of multiple omics data acquired from RNA sequencing,ribosome profiling,and methylated RNA immunoprecipitation sequencing data under STM2457 treatment or METTL3 knockout.We also constructed a model for the regulation of the translation of METTL3 and PD-L1.Finally,we found PD-L1 upregulation by STM2457 in vivo and in vitro.In conclusion,STM2457 is a potential novel suppressor based on its inhibitory effect on tumor progression and may be able to overcome the heterogeneity based on its impact on the translatome.Furthermore,it can improve the immunotherapy outcomes based on PDL1 upregulation in NSCLC.
基金supported by the National Key R&D Program of China(2021YFA1102200,2024YFA1802300 and 2024YFA1107000)The National Natural Science Foundation of China(32225012)+4 种基金Major Project of Guangzhou National Laboratory(GZNL2023A02005)Guangdong Basic and Applied Basic Research Foundation(2025A1515012426,2023A1515010420)Science and Technology Projects in Guangzhou(2023A04J0724)Science and Technology Planning Project of Guangdong Province,China(2023B1212060050,2023B1212120009)Health@InnoHK Program launched by Innovation Technology Commission of the Hong Kong SAR,P.R.China.
文摘N6-methyladenosine(m^(6)A)plays crucial roles in development and cellular reprogramming.During embryonic development,pluripotency transitions from a naïve to a primed state,and modeling the reverse primed-to-naïve transition(PNT)provides a valuable framework for investigating pluripotency regulation.Here,we show that inhibiting METTL3 significantly promotes PNT in an m^(6)A-dependent manner.Mechanistically,we found that suppressing METTL3 and YTHDF2 prolongs the lifetimes of pluripotency-associated mRNAs,such as Nanog and Sox2,during PNT.In addition,Gstp1 was identified as a downstream target of METTL3 inhibition and YTHDF2 knockout.Gstp1 overexpression enhances PNT,whereas its inhibition impedes the transition.Overall,our findings suggest that YTHDF2 facilitates the removal of pluripotency gene transcripts and Gstp1,thereby promoting PNT reprogramming through m^(6)A-mediated posttranscriptional control.
基金supported by grants from the National Natural Science Foundation of China(82202017,32470951,82273214,and 82200119)the Shanghai Science and Technology Commission(22ZR1439600 and 20JC1410100)+2 种基金the Medical Talent Plan of High-level Local Universities in Shanghai(KJ3-0221-22-6338)Chongqing Medical University High-level Talent Introduction and Research Launch Fund(R2062)the Fundamental Research Funds for the Central Universities.
文摘The development and homeostasis of intestinal epithelium are mediated by actively proliferating Lgr5^(+)stem cells,which pos-sess a remarkable self-renewal and differentiation capacity.Recently,our study demonstrated that N^(6)-methyladenosine(m^(6)A)methylation was essential for the survival of colonic stem cells.Here,we show that methyltransferase-like 3(METTL3)expression is downregulated in the colon mucosa in ulcerative colitis(UC)patients and strongly associated with the differentiation and maturation of goblet cells during inflammation.In mice,depletion of Mettl3 significantly inhibits the self-renewal and differentiation of Lgr5^(+)stem cells,especially the differentiation and maturation of goblet cells,resulting in intestinal dysplasia and spontaneous inflammation.Mechanistically,Mettl3 deletion-mediated m^(6)A loss facilitates the expression levels of growth factor receptor binding protein 10(Grb10)and interferon-related developmental regulator 1(Ifrd1)via increasing their messenger RNA stability.We further demonstrate that the levels of GRB10 and IFRD1 are negatively correlated with METTL3 level in UC samples.Collectively,our data indicate that METTL3 enhances the self-renewal and differentiation of Lgr5^(+)stem cells during intestinal development and inflammation,and thus it may be a potential therapeutic target for UC treatment.
基金National Natural Science Foundation of China(31988101 to YGC,92354306 to YL)National Key Research and Development Program of China(2023YFA1800603)+2 种基金Natural Science Foundation of Jiangxi Province(20224ACB209001)Beijing Science and Technology Plan(Z231100007223006)Shenzhen Medical Research Fund(B2302022)to YGC.
文摘Intestinal epithelial cells(IECs)are pivotal for maintaining intestinal homeostasis through self-renewal,proliferation,differentiation,and regulated cell death.While apoptosis and necroptosis are recognized as distinct pathways,their intricate interplay remains elusive.In this study,we report that Mettl3-mediatedm6A modification maintains intesti-nal homeostasis by impeding epithelial cell death.Mettl3 knockout induces both apoptosis and necroptosis in IECs.Targeting different modes of cell death with specific inhibitors unveils that RIPK1 kinase activity is critical for the cell death triggered by Mettl3 knockout.Mechanistically,this occurs via them6A-mediated transcriptional regulation of Atf3,a transcription factor that directly binds to Cflar,the gene encoding the anti-cell death protein cFLIP.cFLIP inhibits RIPK1 activity,thereby suppressing downstream apoptotic and necroptotic signaling.Together,these find-ings delineate the essential role of the METTL3-ATF3-cFLIP axis in homeostatic regulation of the intestinal epithelium by blocking RIPK1 activity.
基金National Natural Science Foundation of China,Grant/Award Numbers:81872409,82173362Natural Science Foundation of Guangdong Province,Grant/Award Numbers:2018A030313610,2019A1515110110,2020A1515010291+1 种基金The Open Funding of the State Key Laboratory of Oral Diseases,Grant/Award Number:SKLOD2021OF02and the use was approved by the Institutional Review Board of the First Affiliated Hospital of Sun Yat-Sen University(2016074).The nude mouse experi-ments performed were approved by the Laboratory Ani-mal Center of Sun Yat-SenUniversity(SYSU-IACUC-2021-000092).The transgenic mouse experiments performed were approved by the Laboratory Animal Center of Sun Yat-Sen University(SYSU-IACUC-2020-000569).
文摘Background:Cancer cells selectively promote the translation of oncogenic tran-scripts to stimulate cancer progression.Although growing evidence has revealed that tRNA modifications and related genes participate in this process,their roles in head and neck squamous cell carcinoma(HNSCC)remain largely unchar-acterized.Here,we sought to investigate the function and mechanisms of the transfer RNA(tRNA)N7-methylguanosine(m'G)modification in regulating the occurrence and development of HNSCC.Methods:Cell lost of-function and gain-of function assays,xenograft models,conditional knockout and knockin mouse models were used to study the physi-ological functions of tRNA m'G modification in HNSCC tumorigenesis.tRNA modification and expression profiling,mRNA translation profiling and res-cue assays were performed to uncover the underlying molecular mechanisms.Single-cell RNA sequencing(scRNA seq)was conducted to explore the tumor microenvironment changes.Results:The tRNA.m7G methyltransferase complex components Methyltransferase-like 1(METTL1)/WD repeat domain 4(WDR4)were upregulated in HNSCC and associated with a poor prognosis.Functionally,METTL1/WDR4 promoted HNSCC progression and metastasis in cell-based and transgenic mouse models.Mechanistically,ablation of METTL1 reduced the m'G levels of 16 tRNAS,inhibiting the translation of a subset of oncogenic transcripts,including genes related to the phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin(PI3K/AKT/mTOR)signaling pathway.In addition,chemical modulators of the PI3K/Akt/mTOR signaling pathway reversed the effects of Mettll in mouse HNSCC.Furthermore,scRNA-seq results revealed that Mettll knockout in mouse tumor cells altered the immune landscape and cell-cell interaction between the tumor and stromal compartment.Conclusions:The tRNA m?G methyltransferase METTLI was found to promote the development and malignancy of HNSCC through regulating global mRNA translation,including the PI3K/AKT/mTOR signaling pathway,and found to alter immune landscape.METTLI could be a promising treatment target for HNSCC patients.
基金Natural Science Foundation of Guangdong Province,Grant/Award Numbers:2018A030313634,2020A1515010214,2021A1515010473Guangdong Basic and Applied Basic Research Foundation,Grant/Award Numbers:2020A1515010214,2018A030313634,2021A1515010473+1 种基金Young Teacher Foundation of Sun Yat-sen University,Grant/Award Number:19ykpy58China Postdoctoral Science Foundation,Grant/Award Number:2020M683087。
文摘Background:Methyltransferase 3(METTL3)-mediated N6-methyladenosine(m^(6)A)RNA modification has been demonstrated to be a potential factor in promoting gastric cancer(GC).METTL3 regulates a series of signaling pathways by modifying various mRNAs.This study aimed to identify novel METTL3-mediated signaling pathways and explored possible targets for use in the clinical setting of gastric cancer.Methods:To investigate the proliferation and metastatic capacity ofGCcell lines with METTL3 knockdown,a xenograft,lung metastasis,and popliteal lymph node metastasis model was used.Them^(6)A-modified RNA immunoprecipitation(Me-RIP)sequence was utilized to explore the target mRNAs of METTL3.Cell counting kit 8 and transwell assays were performed to investigate the promoting function of pre-B cell leukemia homeobox 1(PBX1)and GTP cyclohydrolase 1(GCH1).Western blotting and chromatin immunoprecipitation were employed to confirm the involvement of the METTL3-PBX1-GCH1 axis.ELISA and liquid chromatography-mass spectrometry were used to explore the biological function of tetrahydrobiopterin(BH_(4)).Results:Knockdown of METTL3 suppressed xenograft tumor growth and lung/lymph node metastasis in vivo.Mechanistically,we found that METTL3 combined with and stabilized PBX1 mRNAs.Chromatin immunoprecipitation(ChIP)and further experiments suggested that PBX1 acted as a transcription factor inducing GCH1 expression.Moreover,the METTL3-PBX1-GCH1 axis increased BH_(4)levels in GC cells,thereby promoting tumor progression.Conclusions:This study suggested that METTL3 enzymes promote tumor growth and lung/lymph node metastasis via METTL3-PBX1-GCH1 axis increasing BH_(4)levels in GC.
基金Strategic Priority Research Program of the Chinese Academy of Sciences(Grant No.XDA16030000)the National Key R&D Program(Grant Nos.2017YFA0103803,2018YFA0107703,and 2018YFA0801200)+2 种基金the National Natural Science Foundation of China(Grant Nos.31621004 and 31770872)the Key Research Projects of the Frontier Science of the Chinese Academy of Sciences(Grant Nos.QYZDY-SSW-SMC002 and QYZDB-SSW-SMC022)the Youth Innovation Promotion Association of Chinese Academy of Sciences(Grant No.CAS2018133)
文摘N6-methyladenosine(m^(6)A)is one of the most abundant modifications on m RNAs and plays important roles in various biological processes.The formation of m^(6)A is catalyzed by a methyltransferase complex(MTC)containing a key factor methyltransferase-like 3(Mettl3).However,the functions of Mettl3 and m^(6)A modification in hepatic lipid and glucose metabolism remain unclear.Here,we showed that both Mettl3 expression and m^(6)A level increased in the livers of mice with high fat diet(HFD)-induced metabolic disorders.Overexpression of Mettl3 aggravated HFDinduced liver metabolic disorders and insulin resistance.In contrast,hepatocyte-specific knockout of Mettl3 significantly alleviated HFD-induced metabolic disorders by slowing weight gain,reducing lipid accumulation,and improving insulin sensitivity.Mechanistically,Mettl3 depletion-mediated m^(6)A loss caused extended RNA half-lives of metabolism-related genes,which consequently protected mice against HFD-induced metabolic syndrome.Our findings reveal a critical role of Mettl3-mediated m^(6)A in HFD-induced metabolic disorders and hepatogenous diabetes.
