A 5'-leader,known initially as the 5'-untranslated region,contains multiple isoforms due to alternative splicing(aS)and alternative transcription start site(aTSS).Therefore,a representative 5'-leader is de...A 5'-leader,known initially as the 5'-untranslated region,contains multiple isoforms due to alternative splicing(aS)and alternative transcription start site(aTSS).Therefore,a representative 5'-leader is demanded to examine the embedded RNA regulatory elements in controlling translation efficiency.Here,we develop a ranking algorithm and a deep-learning model to annotate representative 5'-leaders for five plant species.We rank the intra-sample and inter-sample frequency of aS-mediated transcript isoforms using the Kruskal-Wallis test-based algorithm and identify the representative aS-5'-leader.To further assign a representative 5'-end,we train the deep-learning model 5'leaderP to learn aTsS-mediated 5'-end distribution patterns from cap-analysis gene expression data.The model accurately predicts the 5'-end,confirmed experimentally in Arabidopsis and rice.The representative 5'-leader-contained gene models and 5'leaderP can be accessed at RNAirport(http:/www.rnairport.com/leader5P/).The Stage 1 annotation of 5'-leader records 5'-leader diversity and will pave the way to Ribo-Seq open-reading frame annotation,identical to the project recently initiated by human GENCODE.展开更多
Yellow fever virus(YFV) is a re-emerging virus that can cause life-threatening yellow fever disease in humans.Despite the availability of an effective vaccine,little is known about the replication mechanism of YFV,and...Yellow fever virus(YFV) is a re-emerging virus that can cause life-threatening yellow fever disease in humans.Despite the availability of an effective vaccine,little is known about the replication mechanism of YFV,and there are still no available specific anti-YFV medicines.Herein,by introducing the Renilla luciferase gene(Rluc) into an infectious clone of YFV vaccine strain 17 D,we generated a recombinant virus 17 D-Rluc.2 A via reverse genetics approaches.The 17 D-Rluc.2 A had similar plaque morphology and comparable in vitro growth characteristics with its parental strain.Importantly,the reporter luciferase was efficiently expressed in 17 D-Rluc.2 A-infected mammalian and mosquito cells,and there was a good linear correlation between intracellular luciferase expression and extracellular infectious virion reproduction.Furthermore,by a combination of the 17 D-Rluc.2 A reporter virus and selective 2’-hydroxyl acylation analyzed by primer extension(SHAPE)technology,the conserved 5’-SLA element was shown to be essential for YFV replication,highlighting the capability of17 D-R1 uc.2 A in the investigation of YFV replication.At last,we demonstrated that two compounds with distinct anti-viral mechanisms can effectively inhibit the viral propagation in 17 D-Rluc.2 A-infected cells,demonstrating its potential application in the evaluation of anti-viral medicines.Taken together,the 17 D-Rluc.2 A serves as a useful tool for the study of YFV replication and anti-YFV medicine development.展开更多
Induction and mobilization of transposable elements （TEs） following DNA damage or other stresses has been reported in prokaryotes and eukaryotes. Recently it was discovered that eukaryotic TEs are frequently associa...Induction and mobilization of transposable elements （TEs） following DNA damage or other stresses has been reported in prokaryotes and eukaryotes. Recently it was discovered that eukaryotic TEs are frequently associated with long non-coding RNAs （IncRNAs）, many of which are also upregulated by stress. Yet, it is unknown whether DNA damage-induced transcriptional activation of TEs and IncRNAs occurs sporadically or is a synchronized, genome-wide response. Here we investigated the transcriptome of Arabidopsis wild- type （WT） and ataxia telangiectasia mutated （atm） mutant plants 3 h after induction of DNA damage. In WT, expression of 5.2% of the protein-coding genes is 〉 2-fold changed, whereas in atm plants, only 2.6% of these genes are regulated, and the response of genes associated with DNA repair, replication, and cell cy- cle is largely lost. In contrast, only less than 0.6% of TEs and IncRNAs respond to DNA damage in WT plants, and the regulation of 〉95% of them is ATM-dependent. The ATM-downstream factors BRCA1, DRM1, JMJ30, AGO2, and the ATM-independent AGO4 participate in the regulation of individual TEs and IncRNAs. Remarkably, protein-coding genes located adjacent to DNA damage-responsive TEs and IncRNAs are frequently coexpressed, which is consistent with the hypothesis that TEs and IncRNAs located close to genes commonly function as controlling elements.展开更多
To enable diverse functions and precise regulation,an RNA sequence often folds into complex yet distinct structures in different cellular states.Probing RNA in its native environment is essential to uncovering RNA str...To enable diverse functions and precise regulation,an RNA sequence often folds into complex yet distinct structures in different cellular states.Probing RNA in its native environment is essential to uncovering RNA structures of biological contexts.However,current methods generally require large amounts of input RNA and are challenging for physiologically relevant use.Here,we report smartSHAPE,a new RNA structure probing method that requires very low amounts of RNA input due to the largely reduced artefact of probing signals and increased efficiency of library construction.Using smartSHAPE,we showcased the profiling of the RNA structure landscape of mouse intestinal macrophages upon inflammation,and provided evidence that RNA conformational changes regulate immune responses.These results demonstrate that smartSHAPE can greatly expand the scope of RNA structure-based investigations in practical biological systems,and also provide a research paradigm for the study of post-transcriptional regulation.展开更多
基金supported by grants from the National Key R&D Program of China(2023ZD04073)the Major Project of Hubei Hongshan Laboratory(2022hszd016)+1 种基金the Key Research and Development Program of Hubei Province(2022BFE003)the National Natural Science Foundation of China(32070284)to G.Xu.
文摘A 5'-leader,known initially as the 5'-untranslated region,contains multiple isoforms due to alternative splicing(aS)and alternative transcription start site(aTSS).Therefore,a representative 5'-leader is demanded to examine the embedded RNA regulatory elements in controlling translation efficiency.Here,we develop a ranking algorithm and a deep-learning model to annotate representative 5'-leaders for five plant species.We rank the intra-sample and inter-sample frequency of aS-mediated transcript isoforms using the Kruskal-Wallis test-based algorithm and identify the representative aS-5'-leader.To further assign a representative 5'-end,we train the deep-learning model 5'leaderP to learn aTsS-mediated 5'-end distribution patterns from cap-analysis gene expression data.The model accurately predicts the 5'-end,confirmed experimentally in Arabidopsis and rice.The representative 5'-leader-contained gene models and 5'leaderP can be accessed at RNAirport(http:/www.rnairport.com/leader5P/).The Stage 1 annotation of 5'-leader records 5'-leader diversity and will pave the way to Ribo-Seq open-reading frame annotation,identical to the project recently initiated by human GENCODE.
基金This work is supported by the National Natural Science Foundation of China(81871632 and 32070183)the Natural Science Foundation of Guangdong Province(2020A1515010656)+1 种基金the Creative Research Group Foster Project of the Sun Yat-sen Universitysupported by the One-Hundred People Project of the Sun Yat-sen University。
文摘Yellow fever virus(YFV) is a re-emerging virus that can cause life-threatening yellow fever disease in humans.Despite the availability of an effective vaccine,little is known about the replication mechanism of YFV,and there are still no available specific anti-YFV medicines.Herein,by introducing the Renilla luciferase gene(Rluc) into an infectious clone of YFV vaccine strain 17 D,we generated a recombinant virus 17 D-Rluc.2 A via reverse genetics approaches.The 17 D-Rluc.2 A had similar plaque morphology and comparable in vitro growth characteristics with its parental strain.Importantly,the reporter luciferase was efficiently expressed in 17 D-Rluc.2 A-infected mammalian and mosquito cells,and there was a good linear correlation between intracellular luciferase expression and extracellular infectious virion reproduction.Furthermore,by a combination of the 17 D-Rluc.2 A reporter virus and selective 2’-hydroxyl acylation analyzed by primer extension(SHAPE)technology,the conserved 5’-SLA element was shown to be essential for YFV replication,highlighting the capability of17 D-R1 uc.2 A in the investigation of YFV replication.At last,we demonstrated that two compounds with distinct anti-viral mechanisms can effectively inhibit the viral propagation in 17 D-Rluc.2 A-infected cells,demonstrating its potential application in the evaluation of anti-viral medicines.Taken together,the 17 D-Rluc.2 A serves as a useful tool for the study of YFV replication and anti-YFV medicine development.
文摘Induction and mobilization of transposable elements （TEs） following DNA damage or other stresses has been reported in prokaryotes and eukaryotes. Recently it was discovered that eukaryotic TEs are frequently associated with long non-coding RNAs （IncRNAs）, many of which are also upregulated by stress. Yet, it is unknown whether DNA damage-induced transcriptional activation of TEs and IncRNAs occurs sporadically or is a synchronized, genome-wide response. Here we investigated the transcriptome of Arabidopsis wild- type （WT） and ataxia telangiectasia mutated （atm） mutant plants 3 h after induction of DNA damage. In WT, expression of 5.2% of the protein-coding genes is 〉 2-fold changed, whereas in atm plants, only 2.6% of these genes are regulated, and the response of genes associated with DNA repair, replication, and cell cy- cle is largely lost. In contrast, only less than 0.6% of TEs and IncRNAs respond to DNA damage in WT plants, and the regulation of 〉95% of them is ATM-dependent. The ATM-downstream factors BRCA1, DRM1, JMJ30, AGO2, and the ATM-independent AGO4 participate in the regulation of individual TEs and IncRNAs. Remarkably, protein-coding genes located adjacent to DNA damage-responsive TEs and IncRNAs are frequently coexpressed, which is consistent with the hypothesis that TEs and IncRNAs located close to genes commonly function as controlling elements.
基金the National Key R&D Program of China(2019YFA0110002 and 2018YFA0107603 to Q.C.Z,and 2020YFA0509100 to X.H.)National Natural Science Foundation of China(Grants No.32125007,91940306,91740204,and 31761163007 to Q.C.Z,and 31725010,31821003,31991174,32030037,82150105 to X.H.)Research Grants Council of the Hong Kong SAR,China Project No.N_CityU110/17 to C.K.K.
文摘To enable diverse functions and precise regulation,an RNA sequence often folds into complex yet distinct structures in different cellular states.Probing RNA in its native environment is essential to uncovering RNA structures of biological contexts.However,current methods generally require large amounts of input RNA and are challenging for physiologically relevant use.Here,we report smartSHAPE,a new RNA structure probing method that requires very low amounts of RNA input due to the largely reduced artefact of probing signals and increased efficiency of library construction.Using smartSHAPE,we showcased the profiling of the RNA structure landscape of mouse intestinal macrophages upon inflammation,and provided evidence that RNA conformational changes regulate immune responses.These results demonstrate that smartSHAPE can greatly expand the scope of RNA structure-based investigations in practical biological systems,and also provide a research paradigm for the study of post-transcriptional regulation.
