Inherited retinal dystrophies (IRDs) are major causes of visual impairment and irreversible blindness worldwide, while the precise molecular and genetic mechanisms are still elusive. N6-methyladenosine (m^(6)A) modifi...Inherited retinal dystrophies (IRDs) are major causes of visual impairment and irreversible blindness worldwide, while the precise molecular and genetic mechanisms are still elusive. N6-methyladenosine (m^(6)A) modification is the most prevalent internal modification in eukaryotic mRNA. YTH domain containing 2 (YTHDC2), an m^(6)A reader protein, has recently been identified as a key player in germline development and human cancer. However, its contribution to retinal function remains unknown. Here, we explore the role of YTHDC2 in the visual function of retinal rod photoreceptors by generating rod-specific Ythdc2 knockout mice. Results show that Ythdc2 deficiency in rods causes diminished scotopic ERG responses and progressive retinal degeneration. Multi-omics analysis further identifies Ppef2 and Pde6b as the potential targets of YTHDC2 in the retina. Specifically, via its YTH domain, YTHDC2 recognizes and binds m^(6)A-modified Ppef2 mRNA at the coding sequence and Pde6b mRNA at the 5′-UTR, resulting in enhanced translation efficiency without affecting mRNA levels. Compromised translation efficiency of Ppef2 and Pde6b after YTHDC2 depletion ultimately leads to decreased protein levels in the retina, impaired retinal function, and progressive rod death. Collectively, our finding highlights the importance of YTHDC2 in visual function and photoreceptor survival, which provides an unreported elucidation of IRD pathogenesis via epitranscriptomics.展开更多
目的探讨m^(6)A结合蛋白YTHDC2在结直肠癌发展过程中的分子机制及其所参与的信号通路。方法通过The Human Protein Atlas和GEPIA网站分析肿瘤基因组图谱(TCGA)数据库中YTHDC2与结直肠癌的关系,探究相关的信号转导途径和生物学过程。体...目的探讨m^(6)A结合蛋白YTHDC2在结直肠癌发展过程中的分子机制及其所参与的信号通路。方法通过The Human Protein Atlas和GEPIA网站分析肿瘤基因组图谱(TCGA)数据库中YTHDC2与结直肠癌的关系,探究相关的信号转导途径和生物学过程。体外培养结直肠癌HCT116和Caco2细胞系,对YTHDC2进行过表达或敲低后,采用RT-PCR和Western blot检测p38MAPK、p-p38MAPK及其下游凋亡相关蛋白在丝裂原活化蛋白激酶(MAPK)信号通路中的表达,流式细胞术检测细胞凋亡率。结果The Human Protein Atlas和GEPIA分析结果表明,肿瘤组织中YTHDC2表达水平较癌旁组织存在降低趋势,YTHDC2的表达增加与结直肠癌患者总体生存率提高呈正相关。基于cBioPortal和Xena数据库中结直肠癌相关的基因表达量和临床数据以及KEGG和GO数据库中的注释基因集,基因集富集分析(GSEA)提示,YTHDC2具有调控MAPK信号通路的作用。流式细胞术检测结果显示,敲除YTHDC2时细胞凋亡比率显著减少,过表达时则显著增加,Western blot检测结果显示,p38MAPK表达未见显著变化,p-p38MAPK在YTHDC2过表达时显著升高,敲除时显著降低。基因表达谱交互分析(GEPIA)结果提示,凋亡蛋白的表达与YTHDC2的表达呈正相关,与RT-PCR和Western blot检测结果相符合。结论YTHDC2激活p38MAPK信号途径中外源性死亡受体和内源性线粒体凋亡通路调控结直肠癌细胞的凋亡。展开更多
Background Liver ischemia/reperfusion(I/R)injury is usually caused by hepatic inflow occlusion during liver surgery,and is frequently observed during war wounds and trauma.Hepatocyte ferroptosis plays a critical role ...Background Liver ischemia/reperfusion(I/R)injury is usually caused by hepatic inflow occlusion during liver surgery,and is frequently observed during war wounds and trauma.Hepatocyte ferroptosis plays a critical role in liver I/R injury,however,it remains unclear whether this process is controlled or regulated by members of the DEAD/DExH-box helicase(DDX/DHX)family.Methods The expression of DDX/DHX family members during liver I/R injury was screened using transcriptome analysis.Hepatocyte-specific Dhx58 knockout mice were constructed,and a partial liver I/R operation was performed.Single-cell RNA sequencing(scRNA-seq)in the liver post I/R suggested enhanced ferroptosis by Dhx58hep−/−.The mRNAs and proteins associated with DExH-box helicase 58(DHX58)were screened using RNA immunoprecipitation-sequencing(RIP-seq)and IP-mass spectrometry(IP-MS).Results Excessive production of reactive oxygen species(ROS)decreased the expression of the IFN-stimulated gene Dhx58 in hepatocytes and promoted hepatic ferroptosis,while treatment using IFN-αincreased DHX58 expression and prevented ferroptosis during liver I/R injury.Mechanistically,DHX58 with RNA-binding activity constitutively associates with the mRNA of glutathione peroxidase 4(GPX4),a central ferroptosis suppressor,and recruits the m6A reader YT521-B homology domain containing 2(YTHDC2)to promote the translation of Gpx4 mRNA in an m6A-dependent manner,thus enhancing GPX4 protein levels and preventing hepatic ferroptosis.Conclusions This study provides mechanistic evidence that IFN-αstimulates DHX58 to promote the translation of m6A-modified Gpx4 mRNA,suggesting the potential clinical application of IFN-αin the prevention of hepatic ferroptosis during liver I/R injury.展开更多
Background:As a ubiquitous reversible epigenetic RNA modification,N6-methyladenosine(m6A)plays crucial regulatory roles in multiple biological pathways.However,its functional mechanisms in sex determination and differ...Background:As a ubiquitous reversible epigenetic RNA modification,N6-methyladenosine(m6A)plays crucial regulatory roles in multiple biological pathways.However,its functional mechanisms in sex determination and differentiation during gonadal development of chicken embryos are not clear.Therefore,we established a transcriptome-wide m6A map in the female and male chicken left gonads of embryonic day 7(E7)by methylated RNA immunoprecipitation sequencing(MeRIP-seq)to offer insight into the landscape of m6A methylation and investigate the post-transcriptional modification underlying gonadal differentiation.Results:The chicken embryonic gonadal transcriptome was extensively methylated.We found 15,191 and 16,111 m6A peaks in the female and male left gonads,respectively,which were mainly enriched in the coding sequence(CDS)and stop codon.Among these m6A peaks,we identified that 1013 and 751 were hypermethylated in females and males,respectively.These differential peaks covered 281 and 327 genes,such as BMP2,SMAD2,SOX9 and CYP19A1,which were primarily associated with development,morphogenesis and sex differentiation by functional enrichment.Further analysis revealed that the m6A methylation level was positively correlated with gene expression abundance.Furthermore,we found that YTHDC2 could regulate the expression of sex-related genes,especially HEMGN and SOX9,in male mesonephros/gonad mingle cells,which was verified by in vitro experiments,suggesting a regulatory role of m6A methylation in chicken gonad differentiation.Conclusions:This work provided a comprehensive m6A methylation profile of chicken embryonic gonads and revealed YTHDC2 as a key regulator responsible for sex differentiation.Our results contribute to a better understanding of epigenetic factors involved in chicken sex determination and differentiation and to promoting the future development of sex manipulation in poultry industry.展开更多
基金supported by the National Natural Science Foundation of China(81970841,82101160,82121003)the Department of Science and Technology of Sichuan Province(2023ZYD0172,2023YFS0161)+3 种基金the program of Science and Technology International Cooperation Project of Qinghai province(China)(No.2022-HZ-814)Sichuan Intellectual Property Office(China)(No.2022-ZS-0070)the CAMS Innovation Fund for Medical Sciences(2019-12M-5-032)Open Project of Henan Provincial Key Laboratory of Ophthalmology and Visual Science(20KFKT02).
文摘Inherited retinal dystrophies (IRDs) are major causes of visual impairment and irreversible blindness worldwide, while the precise molecular and genetic mechanisms are still elusive. N6-methyladenosine (m^(6)A) modification is the most prevalent internal modification in eukaryotic mRNA. YTH domain containing 2 (YTHDC2), an m^(6)A reader protein, has recently been identified as a key player in germline development and human cancer. However, its contribution to retinal function remains unknown. Here, we explore the role of YTHDC2 in the visual function of retinal rod photoreceptors by generating rod-specific Ythdc2 knockout mice. Results show that Ythdc2 deficiency in rods causes diminished scotopic ERG responses and progressive retinal degeneration. Multi-omics analysis further identifies Ppef2 and Pde6b as the potential targets of YTHDC2 in the retina. Specifically, via its YTH domain, YTHDC2 recognizes and binds m^(6)A-modified Ppef2 mRNA at the coding sequence and Pde6b mRNA at the 5′-UTR, resulting in enhanced translation efficiency without affecting mRNA levels. Compromised translation efficiency of Ppef2 and Pde6b after YTHDC2 depletion ultimately leads to decreased protein levels in the retina, impaired retinal function, and progressive rod death. Collectively, our finding highlights the importance of YTHDC2 in visual function and photoreceptor survival, which provides an unreported elucidation of IRD pathogenesis via epitranscriptomics.
文摘目的探讨m^(6)A结合蛋白YTHDC2在结直肠癌发展过程中的分子机制及其所参与的信号通路。方法通过The Human Protein Atlas和GEPIA网站分析肿瘤基因组图谱(TCGA)数据库中YTHDC2与结直肠癌的关系,探究相关的信号转导途径和生物学过程。体外培养结直肠癌HCT116和Caco2细胞系,对YTHDC2进行过表达或敲低后,采用RT-PCR和Western blot检测p38MAPK、p-p38MAPK及其下游凋亡相关蛋白在丝裂原活化蛋白激酶(MAPK)信号通路中的表达,流式细胞术检测细胞凋亡率。结果The Human Protein Atlas和GEPIA分析结果表明,肿瘤组织中YTHDC2表达水平较癌旁组织存在降低趋势,YTHDC2的表达增加与结直肠癌患者总体生存率提高呈正相关。基于cBioPortal和Xena数据库中结直肠癌相关的基因表达量和临床数据以及KEGG和GO数据库中的注释基因集,基因集富集分析(GSEA)提示,YTHDC2具有调控MAPK信号通路的作用。流式细胞术检测结果显示,敲除YTHDC2时细胞凋亡比率显著减少,过表达时则显著增加,Western blot检测结果显示,p38MAPK表达未见显著变化,p-p38MAPK在YTHDC2过表达时显著升高,敲除时显著降低。基因表达谱交互分析(GEPIA)结果提示,凋亡蛋白的表达与YTHDC2的表达呈正相关,与RT-PCR和Western blot检测结果相符合。结论YTHDC2激活p38MAPK信号途径中外源性死亡受体和内源性线粒体凋亡通路调控结直肠癌细胞的凋亡。
基金National Key Research and Development Program of China(2023YFC2505900)National Natural Science Foundation of China(92269204,82171755,92369106,82171749,82171811,82073184)+1 种基金Military Outstanding Youth Program(2020QN06119,01-SWK JYCJJ07,23SWAQ53)Program of Leading Talents in Shanghai,and Shanghai Shuguang Program(20SG39)。
文摘Background Liver ischemia/reperfusion(I/R)injury is usually caused by hepatic inflow occlusion during liver surgery,and is frequently observed during war wounds and trauma.Hepatocyte ferroptosis plays a critical role in liver I/R injury,however,it remains unclear whether this process is controlled or regulated by members of the DEAD/DExH-box helicase(DDX/DHX)family.Methods The expression of DDX/DHX family members during liver I/R injury was screened using transcriptome analysis.Hepatocyte-specific Dhx58 knockout mice were constructed,and a partial liver I/R operation was performed.Single-cell RNA sequencing(scRNA-seq)in the liver post I/R suggested enhanced ferroptosis by Dhx58hep−/−.The mRNAs and proteins associated with DExH-box helicase 58(DHX58)were screened using RNA immunoprecipitation-sequencing(RIP-seq)and IP-mass spectrometry(IP-MS).Results Excessive production of reactive oxygen species(ROS)decreased the expression of the IFN-stimulated gene Dhx58 in hepatocytes and promoted hepatic ferroptosis,while treatment using IFN-αincreased DHX58 expression and prevented ferroptosis during liver I/R injury.Mechanistically,DHX58 with RNA-binding activity constitutively associates with the mRNA of glutathione peroxidase 4(GPX4),a central ferroptosis suppressor,and recruits the m6A reader YT521-B homology domain containing 2(YTHDC2)to promote the translation of Gpx4 mRNA in an m6A-dependent manner,thus enhancing GPX4 protein levels and preventing hepatic ferroptosis.Conclusions This study provides mechanistic evidence that IFN-αstimulates DHX58 to promote the translation of m6A-modified Gpx4 mRNA,suggesting the potential clinical application of IFN-αin the prevention of hepatic ferroptosis during liver I/R injury.
基金funded in part by grants from China Agricultural Research System(CARS-40).
文摘Background:As a ubiquitous reversible epigenetic RNA modification,N6-methyladenosine(m6A)plays crucial regulatory roles in multiple biological pathways.However,its functional mechanisms in sex determination and differentiation during gonadal development of chicken embryos are not clear.Therefore,we established a transcriptome-wide m6A map in the female and male chicken left gonads of embryonic day 7(E7)by methylated RNA immunoprecipitation sequencing(MeRIP-seq)to offer insight into the landscape of m6A methylation and investigate the post-transcriptional modification underlying gonadal differentiation.Results:The chicken embryonic gonadal transcriptome was extensively methylated.We found 15,191 and 16,111 m6A peaks in the female and male left gonads,respectively,which were mainly enriched in the coding sequence(CDS)and stop codon.Among these m6A peaks,we identified that 1013 and 751 were hypermethylated in females and males,respectively.These differential peaks covered 281 and 327 genes,such as BMP2,SMAD2,SOX9 and CYP19A1,which were primarily associated with development,morphogenesis and sex differentiation by functional enrichment.Further analysis revealed that the m6A methylation level was positively correlated with gene expression abundance.Furthermore,we found that YTHDC2 could regulate the expression of sex-related genes,especially HEMGN and SOX9,in male mesonephros/gonad mingle cells,which was verified by in vitro experiments,suggesting a regulatory role of m6A methylation in chicken gonad differentiation.Conclusions:This work provided a comprehensive m6A methylation profile of chicken embryonic gonads and revealed YTHDC2 as a key regulator responsible for sex differentiation.Our results contribute to a better understanding of epigenetic factors involved in chicken sex determination and differentiation and to promoting the future development of sex manipulation in poultry industry.
