Researchers commonly use cyclization recombination enzyme/locus of X-over P1(Cre/loxP)technology-based conditional gene knockouts of model mice to investigate the functional roles of genes of interest in Sertoli and L...Researchers commonly use cyclization recombination enzyme/locus of X-over P1(Cre/loxP)technology-based conditional gene knockouts of model mice to investigate the functional roles of genes of interest in Sertoli and Leydig cells within the testis.However,the shortcomings of these genetic tools include high costs,lengthy experimental periods,and limited accessibility for researchers.Therefore,exploring alternative gene silencing techniques is of great practical value.In this study,we employed adeno-associated virus(AAV)as a vector for gene silencing in Sertoli and Leydig cells.Our findings demonstrated that AAV serotypes 1,8,and 9 exhibited high infection efficiency in both types of testis cells.Importantly,we discovered that all three AAV serotypes exhibited exquisite specificity in targeting Sertoli cells via tubular injection while demonstrating remarkable selectivity in targeting Leydig cells via interstitial injection.We achieved cell-specific knockouts of the steroidogenic acute regulatory(Star)and luteinizing hormone/human chorionic gonadotropin receptor(Lhcgr)genes in Leydig cells,but not in Sertoli cells,using AAV9-single guide RNA(sgRNA)-mediated gene editing in Rosa26-LSL-Cas9mice.Knockdown of androgen receptor(Ar)gene expression in Sertoli cells of wild-type mice was achieved via tubular injection of AAV9-short hairpinRNA(shRNA)-mediated targeting.Our findings offer technical approaches for investigating gene function in Sertoli and Leydig cells through AAV9-mediated gene silencing.展开更多
In the article“Silencing of the long non-coding RNA LINC00265 triggers autophagy and apoptosis in lung cancer by reducing protein stability of SIN3A oncogene”(Oncology Research.2024,Vol.32,No.7,pp.1185–1195.doi:10....In the article“Silencing of the long non-coding RNA LINC00265 triggers autophagy and apoptosis in lung cancer by reducing protein stability of SIN3A oncogene”(Oncology Research.2024,Vol.32,No.7,pp.1185–1195.doi:10.32604/or.2023.030771,https://www.techscience.com/or/v32n7/57163),an inadvertent error occurred during the compilation of Fig.3H.This needed corrections to ensure the accuracy and integrity of the data presented.展开更多
The occurrence,development,and metastasis of tumors often entail abnormal expression of genetic substances.Monitoring and regulating changes in intracellular nucleic acid substances hold promise for achieving accurate...The occurrence,development,and metastasis of tumors often entail abnormal expression of genetic substances.Monitoring and regulating changes in intracellular nucleic acid substances hold promise for achieving accurate tumor diagnosis and effective treatment.However,the effectiveness of integrated tumor diagnosis and treatment based on functional nucleic acids still needs to be improved.In this study,we engineered a multifunctional nucleic acid delivery system grounded in a cationic covalent organic framework carrier.This system not only showcases effective gene silencing but also boasts high sensitivity in detecting miR21 levels within tumor cells,enabling real-time monitoring of tumor gene therapy efficacy.The construction of this integrated functional nucleic acid delivery platform provides new ideas for precise tumor detection and effective tumor treatment.展开更多
Since transgene silencing was found in transgenic plants,many scholars have studied it extensively and considered that it has three functional mechanisms:post dependent gene silencing,transcriptional gene silencing,p...Since transgene silencing was found in transgenic plants,many scholars have studied it extensively and considered that it has three functional mechanisms:post dependent gene silencing,transcriptional gene silencing,post transcriptional gene silencing.At the moment,people have mainly focused on the study of post transcriptional gene silencing and found its features:extensivity,conduction and peculiarity,also put forward some hypothesis for its mechanisms,for example,RNA threshold model,aberrant RNA model,inter or intra molecular base pairing model and so on.Furthermore,post transcriptional gene silencing is being applied in gene engineering of plants.Recently the people have found that post transcriptional gene silencing has bearing on capacity plants resisting virus.Many researchers have studied post transcriptional gene silencing,but there are some questions which need be solved in the future.This article summarizes progresses in features,mechanisms,applies of post transcriptional gene silencing about transgenic plants.展开更多
[Objective]The aim was to compare the silencing effect of three vectors containing shRNA quartet of different lengths at cellular level and individual level.[Method]To silence EGFP gene,three shRNA quartets of 21,27 a...[Objective]The aim was to compare the silencing effect of three vectors containing shRNA quartet of different lengths at cellular level and individual level.[Method]To silence EGFP gene,three shRNA quartets of 21,27 and 29 bp composed of hU6,mU6,h7SK and hH1 promoters were constructed and then cloned into silencing vectors.After the vectors were transfected into Vero cells or injected into mouse muscle,real-time fluorescence quantitative PCR was conducted to detect the mRNA level of EGFP gene.[Result]All the three vectors showed better silencing effect;in which,the silencing effect at the cellular level was far higher than that at individual level.[Conclusion]Multi-site tandem expression shRNA may show better silencing effect;intramuscular injection maybe a transgenic method for baby mouse.展开更多
Rice blast disease is one of the most devastating diseases in rice production,which severely affects the high and stable yield of rice.The formation of appressorium plays a key role in the pathogenesis of Magnaporthe ...Rice blast disease is one of the most devastating diseases in rice production,which severely affects the high and stable yield of rice.The formation of appressorium plays a key role in the pathogenesis of Magnaporthe grisea in rice.It has been confirmed that a P-type ATPase （P-ATPase） is involved in the formation of appressorium.A number of small molecular substances are able to enter the pathogen from the host during the interactions between pathogens and hosts,thus resisting the infection of pathogens.In this study,a 232 bp DNA sequence with good specificity from the first exon of P-ATPase gene MgAPT2 was used as an interference fragment and was inserted into interference vector forward and reversely.The interfering vector was then transformed into rice blast-susceptible rice variety Nipponbare via Agrobacterium-mediated transformation.Identification of rice plants inoculated with M.grisea at the seedling stage and detection of the expression level of P-ATPase gene MgAPT2 showed that the expression level of MgAPT2 gene in transgenic plants was reduced and the rice blast resistance was improved.This study provided a new way for the innovation of rice germplasm resources resistant to rice blast disease.展开更多
As a large family of RNA helicases,DEAD-box(DDX)RNA helicases play crucial roles in almost all cellular RNA processing activities.However,the role of the DDX gene family in cold tolerance of mei(Prunus mume)remains un...As a large family of RNA helicases,DEAD-box(DDX)RNA helicases play crucial roles in almost all cellular RNA processing activities.However,the role of the DDX gene family in cold tolerance of mei(Prunus mume)remains unclear.In this study,we identified 45 DDX genes through whole-genome analysis unevenly distributed across eight chromosomes and scaffolds of mei.Based on the phylogenetic tree and gene structure analysis,the DDX genes were classified into nine subfamilies based on their motif compositions and intron-exon structures.The results of synteny analysis showed that segmental duplication was considered a major factor contributing to the amplification of the PmDDX family.RNA-Seq and qRT-PCR results revealed differential expression of PmDDX genes under cold stress.Among these,PmDDX39 was significantly up-regulated under cold stress,suggesting its positive role in modulating mei cold tolerance.We found that silenced PmDDX39 under cold stress led to greater damage than the wild seedlings via virus-induced gene silencing(VIGS).Conversely,overexpression of PmDDX39 in Arabidopsis enhanced cold stress tolerance.Moreover,dual luciferase and yeast one-hybrid(Y1H)demonstrated that PmDDX39 directly activates the expression of the C-repeat binding factor(PmCBFf)by binding to its promoters.This study provides new insights into the structure,evolution,and functional role of the PmDDX gene family in mei responses to cold stress.展开更多
Virus-induced gene silencing (VIGS) is a recently developed technique for characterizing the function of plant genes by gene transcript suppression and is increasingly used to generate transient loss-of-function assay...Virus-induced gene silencing (VIGS) is a recently developed technique for characterizing the function of plant genes by gene transcript suppression and is increasingly used to generate transient loss-of-function assays. Here we report that the 2mDNA1, a geminivirus satellite vector, can induce efficient gene silencing in Nicotiana tabacum with Tobacco curly shoot virus. We have successfully silenced the β-glucuronidase (GUS) gene in GUS transgenic N. tabacum plants and the sulphur desaturase (Su) gene in five different N. tabacum cultivars. These pronounced and severe silencing phenotypes are persistent and ubiquitous. Once initiated in seedlings, the silencing phenotype lasted for the entire life span of the plants and silencing could be induced in a variety of tissues and organs including leaf, shoot, stem, root, and flower, and achieved at any growth stage. This system works well between 18-32°C. We also silenced the NtEDS1 gene and demonstrated that NtEDS1 is essential for N gene mediated resistance against Tobacco mosaic virus in N. tabacum. The above results indicate that this system has great potential as a versatile VIGS system for routine functional analysis of genes in N. tabacum.展开更多
We hypothesized that RNA interference to silence Nogo-66 receptor gene expression in bone marrow mesenchymal stem cells before transplantation might further improve neurological function in rats with spinal cord trans...We hypothesized that RNA interference to silence Nogo-66 receptor gene expression in bone marrow mesenchymal stem cells before transplantation might further improve neurological function in rats with spinal cord transection injury. After 2 weeks, the number of neurons and BrdU-positive cells in the Nogo-66 receptor gene silencing group was higher than in the bone marrow mesenchymal stem cell group, and significantly greater compared with the model group. After 4 weeks, behavioral performance was signiifcantly enhanced in the model group. Af-ter 8 weeks, the number of horseradish peroxidase-labeled nerve ifbers was higher in the Nogo-66 receptor gene silencing group than in the bone marrow mesenchymal stem cell group, and signiifcantly higher than in the model group. The newly formed nerve ifbers and myelinated ner ve ifbers were detectable in the central transverse plane section in the bone marrow mesenchymal stem cell group and in the Nogo-66 receptor gene silencing group.展开更多
The sea cucumber, Apostichopusjaponicus, undergoes aestivation to improve survival during periods of high-temperature. During aestivation, the metabolic rate is depressed to reduce the consumption of reserved energy. ...The sea cucumber, Apostichopusjaponicus, undergoes aestivation to improve survival during periods of high-temperature. During aestivation, the metabolic rate is depressed to reduce the consumption of reserved energy. We evaluated the role of epigenetic modification on global gene silencing during metabolic rate depression in the sea cucumber. We compared the expression of epigenetic modifiers in active and aestivating sea cucumbers. The expression of three genes involved in DNA methylation and chromatin remodeling （DNA （cytosine-5）-methyltransferase l, Methyl-CpG-binding domain protein 2）, and Chromodomain-helicase-DNA-binding protein 5） was significantly higher during aestivation （Days 20 and 40）. Similarly, we observed an increase in the expression of genes involved in histone acetylation （Histone deacetylase 3） and Histone-binding protein RBBP4） during the early （Days 5 and 10） and late phases （Days 20 and 40） of aestivation. There was no change in the expression of KAT2B, a histone acetyltransferase. However, the expression of histone methylation associated modifiers （Histone-arginine methyltransferase CARMER and Histone-lysine N-methyltransferase MLL5） was significantly higher after 5 d in the aestivating group. The results suggest that the expression of epigenetic modifiers involved in DNA methylation, chromatin remodeling, histone acetylation, and histone methylation is upregulated during aestivation. We hypothesize that these changes regulate global gene silencing during aestivation in A. japonicus.展开更多
Phytic acid is the principal storage form of phosphorus in plant seeds and an essential signalling molecule in several regulatory processes of plant development.However,it is known as an anti-nutrient compound owing t...Phytic acid is the principal storage form of phosphorus in plant seeds and an essential signalling molecule in several regulatory processes of plant development.However,it is known as an anti-nutrient compound owing to its potent chelating property.Thus,reducing the phytic acid content in crops is desirable.Studies involving regulation of MIPS and IPK1 genes to generate low phytate rice have been reported earlier.However,the functional significance of OsITPK and the effect of its down-regulation on phytic acid content and the associated pleiotropic effects on rice have not yet been investigated.In this study,tissue specific RNA interference(RNAi)-mediated down-regulation of a major ITPK homolog(OsITP5/6K-1)resulted in 46.2%decrease in phytic acid content of T2 transgenic seeds with a subsequent 3-fold enhancement in the inorganic phosphorus content.Silencing of OsITP5/6K-1 altered the transcript levels of essential phytic acid pathway genes,without significantly affecting the transcript levels of other OsITPK homologs.Furthermore,the mapping of elements through X-ray microfluorescence analysis revealed significant changes in the spatial distribution pattern and translocation of elements in low phytate seeds.Additionally,low phytate polished seeds exhibited 1.3-fold and 1.6-fold enhancement in iron and zinc content in the grain endosperm,respectively.Silencing of OsITP5/6K-1 also altered the amino acid and myo-inositol content of the transgenic seeds.Our results successfully established that RNAi-mediated silencing of OsITP5/6K-1 gene significantly reduced the phytate levels in seeds without hampering the germination potential of seeds and plant growth.The present study provided an insight into the mechanism of phytic acid biosynthesis pathway.展开更多
To understand the functional identification of large-scale genomic sequences in Forsythia,tobacco rattle virus(TRV)-mediated virus-induced gene silencing(VIGS),suitable for the plant,was explored in this study.The res...To understand the functional identification of large-scale genomic sequences in Forsythia,tobacco rattle virus(TRV)-mediated virus-induced gene silencing(VIGS),suitable for the plant,was explored in this study.The results showed that the TRV-mediated VIGS system could be successfully used in Forsythia for silencing the reporter gene FsPDS(Forsythia phytoene desaturase)using stem infiltration and leaf infiltrationmethods.All the treated plants were pruned below the injection site after 7–15 d infection;the FsPDS was silenced and typical photobleaching symptoms were observed in newly sprouted leaves at the whole-plant level.Meanwhile,this system has been successfully tested and verified through virus detection and qRT-PCR analysis.After the optimization,Forsythia magnesium chelatase subunit H(FsChlH)was silenced successfully in Forsythia using this system,resulting in yellow leaveswith decreased chlorophyll content.The system was stable,highly efficient and had greater rapidity and convenience,which made it suitable to study the function of genes related to physiological pathways such as growth and development,and metabolic regulation in Forsythia.展开更多
Invasive nucleic acids such as transposons and viruses usually exhibit aberrant characteristics,e.g.,unpaired DNA or abnormal doublestranded RNA.Organisms employ a variety of strategies to defend themselves by disting...Invasive nucleic acids such as transposons and viruses usually exhibit aberrant characteristics,e.g.,unpaired DNA or abnormal doublestranded RNA.Organisms employ a variety of strategies to defend themselves by distinguishing self and nonself substances and disabling these invasive nucleic acids.Furthermore,they have developed ways to remember this exposure to invaders and transmit the experience to their descendants.The mechanism underlying this inheritance has remained elusive.Recent research has shed light on the initiation and maintenance of RNA-mediated inherited gene silencing.Small regulatory RNAs play a variety of crucial roles in organisms,including gene regulation,developmental timing,antiviral defense,and genome integrity,via a process termed as RNA interference（RNAi）.Recent research has revealed that small RNAs and the RNAi machinery are engaged in establishing and promoting transgenerational gene silencing.Small RNAs direct the RNAi and chromatin modification machinery to the cognate nucleic acids to regulate gene expression and epigenetic alterations.Notably,these acquired small RNAs and epigenetic changes persist and are transmitted from parents to offspring for multiple generations.Thus,RNAi is a vital determinant of the inheritance of gene silencing and acts as a driving force of evolution.展开更多
Global prophylactic vaccination programmes have helped to curb new hepatitis B virus(HBV)infections.However,it is estimated that nearly 300 million people are chronically infected and have a high risk of developing he...Global prophylactic vaccination programmes have helped to curb new hepatitis B virus(HBV)infections.However,it is estimated that nearly 300 million people are chronically infected and have a high risk of developing hepatocellular carcinoma.As such,HBV remains a serious health priority and the development of novel curative therapeutics is urgently needed.Chronic HBV infection has been attributed to the persistence of the covalently closed circular DNA(cccDNA)which establishes itself as a minichromosome in the nucleus of hepatocytes.As the viral transcription intermediate,the cccDNA is responsible for producing new virions and perpetuating infection.HBV is dependent on various host factors for cccDNA formation and the minichromosome is amenable to epigenetic modifications.Two HBV proteins,X(HBx)and core(HBc)promote viral replication by modulating the cccDNA epigenome and regulating host cell responses.This includes viral and host gene expression,chromatin remodeling,DNA methylation,the antiviral immune response,apoptosis,and ubiquitination.Elimination of the cccDNA minichromosome would result in a sterilizing cure;however,this may be difficult to achieve.Epigenetic therapies could permanently silence the cccDNA minichromosome and promote a functional cure.This review explores the cccDNA epigenome,how host and viral factors influence transcription,and the recent epigenetic therapies and epigenome engineering approaches that have been described.展开更多
基金supported by the National Natural Science Foundation of China(No.82070872 and No.82370854 to JXL)Innovative and Entrepreneurial Team of Jiangsu Province(No.JSSCTD2021 to JXL)+1 种基金China Postdoctoral Science Foundation(2023M741790 to JP)Jiangsu Funding Program for Excellent Postdoctoral Talent(2023ZB558 to JP).
文摘Researchers commonly use cyclization recombination enzyme/locus of X-over P1(Cre/loxP)technology-based conditional gene knockouts of model mice to investigate the functional roles of genes of interest in Sertoli and Leydig cells within the testis.However,the shortcomings of these genetic tools include high costs,lengthy experimental periods,and limited accessibility for researchers.Therefore,exploring alternative gene silencing techniques is of great practical value.In this study,we employed adeno-associated virus(AAV)as a vector for gene silencing in Sertoli and Leydig cells.Our findings demonstrated that AAV serotypes 1,8,and 9 exhibited high infection efficiency in both types of testis cells.Importantly,we discovered that all three AAV serotypes exhibited exquisite specificity in targeting Sertoli cells via tubular injection while demonstrating remarkable selectivity in targeting Leydig cells via interstitial injection.We achieved cell-specific knockouts of the steroidogenic acute regulatory(Star)and luteinizing hormone/human chorionic gonadotropin receptor(Lhcgr)genes in Leydig cells,but not in Sertoli cells,using AAV9-single guide RNA(sgRNA)-mediated gene editing in Rosa26-LSL-Cas9mice.Knockdown of androgen receptor(Ar)gene expression in Sertoli cells of wild-type mice was achieved via tubular injection of AAV9-short hairpinRNA(shRNA)-mediated targeting.Our findings offer technical approaches for investigating gene function in Sertoli and Leydig cells through AAV9-mediated gene silencing.
文摘In the article“Silencing of the long non-coding RNA LINC00265 triggers autophagy and apoptosis in lung cancer by reducing protein stability of SIN3A oncogene”(Oncology Research.2024,Vol.32,No.7,pp.1185–1195.doi:10.32604/or.2023.030771,https://www.techscience.com/or/v32n7/57163),an inadvertent error occurred during the compilation of Fig.3H.This needed corrections to ensure the accuracy and integrity of the data presented.
基金the National Key Research and Development Program of China(No.2021YFB3800900)National Natural Science Foundation of China(No.51925305)the talent cultivation project Funds for the Innovation Laboratory for Sciences and Technologies of Energy Materials of Fujian Province(No.HRTP[2022]52)。
文摘The occurrence,development,and metastasis of tumors often entail abnormal expression of genetic substances.Monitoring and regulating changes in intracellular nucleic acid substances hold promise for achieving accurate tumor diagnosis and effective treatment.However,the effectiveness of integrated tumor diagnosis and treatment based on functional nucleic acids still needs to be improved.In this study,we engineered a multifunctional nucleic acid delivery system grounded in a cationic covalent organic framework carrier.This system not only showcases effective gene silencing but also boasts high sensitivity in detecting miR21 levels within tumor cells,enabling real-time monitoring of tumor gene therapy efficacy.The construction of this integrated functional nucleic acid delivery platform provides new ideas for precise tumor detection and effective tumor treatment.
文摘Since transgene silencing was found in transgenic plants,many scholars have studied it extensively and considered that it has three functional mechanisms:post dependent gene silencing,transcriptional gene silencing,post transcriptional gene silencing.At the moment,people have mainly focused on the study of post transcriptional gene silencing and found its features:extensivity,conduction and peculiarity,also put forward some hypothesis for its mechanisms,for example,RNA threshold model,aberrant RNA model,inter or intra molecular base pairing model and so on.Furthermore,post transcriptional gene silencing is being applied in gene engineering of plants.Recently the people have found that post transcriptional gene silencing has bearing on capacity plants resisting virus.Many researchers have studied post transcriptional gene silencing,but there are some questions which need be solved in the future.This article summarizes progresses in features,mechanisms,applies of post transcriptional gene silencing about transgenic plants.
基金Supported by the National Natural Science Foundation of China(30671495)Project for Breeding New Varieties(2008ZX08006-003)+1 种基金Project for Breeding New Varieties(2009ZX08011-030B)International Cooperation Project(2009BFA012)~~
文摘[Objective]The aim was to compare the silencing effect of three vectors containing shRNA quartet of different lengths at cellular level and individual level.[Method]To silence EGFP gene,three shRNA quartets of 21,27 and 29 bp composed of hU6,mU6,h7SK and hH1 promoters were constructed and then cloned into silencing vectors.After the vectors were transfected into Vero cells or injected into mouse muscle,real-time fluorescence quantitative PCR was conducted to detect the mRNA level of EGFP gene.[Result]All the three vectors showed better silencing effect;in which,the silencing effect at the cellular level was far higher than that at individual level.[Conclusion]Multi-site tandem expression shRNA may show better silencing effect;intramuscular injection maybe a transgenic method for baby mouse.
基金Supported by International Cooperation Project of Jiangsu Province(BZ2011039)Agricultural Technology Independent Innovation Fund of Jiangsu Province[CX(12)2024]~~
文摘Rice blast disease is one of the most devastating diseases in rice production,which severely affects the high and stable yield of rice.The formation of appressorium plays a key role in the pathogenesis of Magnaporthe grisea in rice.It has been confirmed that a P-type ATPase （P-ATPase） is involved in the formation of appressorium.A number of small molecular substances are able to enter the pathogen from the host during the interactions between pathogens and hosts,thus resisting the infection of pathogens.In this study,a 232 bp DNA sequence with good specificity from the first exon of P-ATPase gene MgAPT2 was used as an interference fragment and was inserted into interference vector forward and reversely.The interfering vector was then transformed into rice blast-susceptible rice variety Nipponbare via Agrobacterium-mediated transformation.Identification of rice plants inoculated with M.grisea at the seedling stage and detection of the expression level of P-ATPase gene MgAPT2 showed that the expression level of MgAPT2 gene in transgenic plants was reduced and the rice blast resistance was improved.This study provided a new way for the innovation of rice germplasm resources resistant to rice blast disease.
基金supported by the Fundamental Research Funds for the Central Universities(Grant No.QNTD202503)Forestry and Grassland Science and Technology Innovation Youth Top Talent Project of China(Grant No.2020132608)Beijing High-Precision Discipline Project,Discipline of Ecological Environment of Urban and Rural Human Settlements.
文摘As a large family of RNA helicases,DEAD-box(DDX)RNA helicases play crucial roles in almost all cellular RNA processing activities.However,the role of the DDX gene family in cold tolerance of mei(Prunus mume)remains unclear.In this study,we identified 45 DDX genes through whole-genome analysis unevenly distributed across eight chromosomes and scaffolds of mei.Based on the phylogenetic tree and gene structure analysis,the DDX genes were classified into nine subfamilies based on their motif compositions and intron-exon structures.The results of synteny analysis showed that segmental duplication was considered a major factor contributing to the amplification of the PmDDX family.RNA-Seq and qRT-PCR results revealed differential expression of PmDDX genes under cold stress.Among these,PmDDX39 was significantly up-regulated under cold stress,suggesting its positive role in modulating mei cold tolerance.We found that silenced PmDDX39 under cold stress led to greater damage than the wild seedlings via virus-induced gene silencing(VIGS).Conversely,overexpression of PmDDX39 in Arabidopsis enhanced cold stress tolerance.Moreover,dual luciferase and yeast one-hybrid(Y1H)demonstrated that PmDDX39 directly activates the expression of the C-repeat binding factor(PmCBFf)by binding to its promoters.This study provides new insights into the structure,evolution,and functional role of the PmDDX gene family in mei responses to cold stress.
基金supported by the National Science and Technology Major Projects of China (No. 2009ZX08009-026B)the China Postdoctoral Science Foundation (No. 20090461375)the National Basic Research Program (973) of China (No. 2006CB101903)
文摘Virus-induced gene silencing (VIGS) is a recently developed technique for characterizing the function of plant genes by gene transcript suppression and is increasingly used to generate transient loss-of-function assays. Here we report that the 2mDNA1, a geminivirus satellite vector, can induce efficient gene silencing in Nicotiana tabacum with Tobacco curly shoot virus. We have successfully silenced the β-glucuronidase (GUS) gene in GUS transgenic N. tabacum plants and the sulphur desaturase (Su) gene in five different N. tabacum cultivars. These pronounced and severe silencing phenotypes are persistent and ubiquitous. Once initiated in seedlings, the silencing phenotype lasted for the entire life span of the plants and silencing could be induced in a variety of tissues and organs including leaf, shoot, stem, root, and flower, and achieved at any growth stage. This system works well between 18-32°C. We also silenced the NtEDS1 gene and demonstrated that NtEDS1 is essential for N gene mediated resistance against Tobacco mosaic virus in N. tabacum. The above results indicate that this system has great potential as a versatile VIGS system for routine functional analysis of genes in N. tabacum.
文摘We hypothesized that RNA interference to silence Nogo-66 receptor gene expression in bone marrow mesenchymal stem cells before transplantation might further improve neurological function in rats with spinal cord transection injury. After 2 weeks, the number of neurons and BrdU-positive cells in the Nogo-66 receptor gene silencing group was higher than in the bone marrow mesenchymal stem cell group, and significantly greater compared with the model group. After 4 weeks, behavioral performance was signiifcantly enhanced in the model group. Af-ter 8 weeks, the number of horseradish peroxidase-labeled nerve ifbers was higher in the Nogo-66 receptor gene silencing group than in the bone marrow mesenchymal stem cell group, and signiifcantly higher than in the model group. The newly formed nerve ifbers and myelinated ner ve ifbers were detectable in the central transverse plane section in the bone marrow mesenchymal stem cell group and in the Nogo-66 receptor gene silencing group.
基金Supported by the National Key Technology Research and Development Program(No.2006AA10A411)the Agricultural Seed Project of Shandong Province
文摘The sea cucumber, Apostichopusjaponicus, undergoes aestivation to improve survival during periods of high-temperature. During aestivation, the metabolic rate is depressed to reduce the consumption of reserved energy. We evaluated the role of epigenetic modification on global gene silencing during metabolic rate depression in the sea cucumber. We compared the expression of epigenetic modifiers in active and aestivating sea cucumbers. The expression of three genes involved in DNA methylation and chromatin remodeling （DNA （cytosine-5）-methyltransferase l, Methyl-CpG-binding domain protein 2）, and Chromodomain-helicase-DNA-binding protein 5） was significantly higher during aestivation （Days 20 and 40）. Similarly, we observed an increase in the expression of genes involved in histone acetylation （Histone deacetylase 3） and Histone-binding protein RBBP4） during the early （Days 5 and 10） and late phases （Days 20 and 40） of aestivation. There was no change in the expression of KAT2B, a histone acetyltransferase. However, the expression of histone methylation associated modifiers （Histone-arginine methyltransferase CARMER and Histone-lysine N-methyltransferase MLL5） was significantly higher after 5 d in the aestivating group. The results suggest that the expression of epigenetic modifiers involved in DNA methylation, chromatin remodeling, histone acetylation, and histone methylation is upregulated during aestivation. We hypothesize that these changes regulate global gene silencing during aestivation in A. japonicus.
文摘Phytic acid is the principal storage form of phosphorus in plant seeds and an essential signalling molecule in several regulatory processes of plant development.However,it is known as an anti-nutrient compound owing to its potent chelating property.Thus,reducing the phytic acid content in crops is desirable.Studies involving regulation of MIPS and IPK1 genes to generate low phytate rice have been reported earlier.However,the functional significance of OsITPK and the effect of its down-regulation on phytic acid content and the associated pleiotropic effects on rice have not yet been investigated.In this study,tissue specific RNA interference(RNAi)-mediated down-regulation of a major ITPK homolog(OsITP5/6K-1)resulted in 46.2%decrease in phytic acid content of T2 transgenic seeds with a subsequent 3-fold enhancement in the inorganic phosphorus content.Silencing of OsITP5/6K-1 altered the transcript levels of essential phytic acid pathway genes,without significantly affecting the transcript levels of other OsITPK homologs.Furthermore,the mapping of elements through X-ray microfluorescence analysis revealed significant changes in the spatial distribution pattern and translocation of elements in low phytate seeds.Additionally,low phytate polished seeds exhibited 1.3-fold and 1.6-fold enhancement in iron and zinc content in the grain endosperm,respectively.Silencing of OsITP5/6K-1 also altered the amino acid and myo-inositol content of the transgenic seeds.Our results successfully established that RNAi-mediated silencing of OsITP5/6K-1 gene significantly reduced the phytate levels in seeds without hampering the germination potential of seeds and plant growth.The present study provided an insight into the mechanism of phytic acid biosynthesis pathway.
基金Thanks for the technical support of Dr.Daqi Fu and Dr.Lanhuan Meng of China Agricultural University.This work was supported by Beijing Municipal Science and Technology Project(Grant No.Z181100002418006)the Fundamental Research Fund for the Central University(Grant No.2015ZCQ-YL-03)the World-Class Discipline Construction and Characteristic Development Guidance Funds for Beijing Forestry University(Grant No.2019XKJS0323).
文摘To understand the functional identification of large-scale genomic sequences in Forsythia,tobacco rattle virus(TRV)-mediated virus-induced gene silencing(VIGS),suitable for the plant,was explored in this study.The results showed that the TRV-mediated VIGS system could be successfully used in Forsythia for silencing the reporter gene FsPDS(Forsythia phytoene desaturase)using stem infiltration and leaf infiltrationmethods.All the treated plants were pruned below the injection site after 7–15 d infection;the FsPDS was silenced and typical photobleaching symptoms were observed in newly sprouted leaves at the whole-plant level.Meanwhile,this system has been successfully tested and verified through virus detection and qRT-PCR analysis.After the optimization,Forsythia magnesium chelatase subunit H(FsChlH)was silenced successfully in Forsythia using this system,resulting in yellow leaveswith decreased chlorophyll content.The system was stable,highly efficient and had greater rapidity and convenience,which made it suitable to study the function of genes related to physiological pathways such as growth and development,and metabolic regulation in Forsythia.
基金supported by the grants from the State Key Development Program for Basic Research of China (No.2011 CBA01103)the National Natural Science Foundation of China(No.31171254)the Fundamental Research Funds for Central Universities(No.WK 2060190018)
文摘Invasive nucleic acids such as transposons and viruses usually exhibit aberrant characteristics,e.g.,unpaired DNA or abnormal doublestranded RNA.Organisms employ a variety of strategies to defend themselves by distinguishing self and nonself substances and disabling these invasive nucleic acids.Furthermore,they have developed ways to remember this exposure to invaders and transmit the experience to their descendants.The mechanism underlying this inheritance has remained elusive.Recent research has shed light on the initiation and maintenance of RNA-mediated inherited gene silencing.Small regulatory RNAs play a variety of crucial roles in organisms,including gene regulation,developmental timing,antiviral defense,and genome integrity,via a process termed as RNA interference（RNAi）.Recent research has revealed that small RNAs and the RNAi machinery are engaged in establishing and promoting transgenerational gene silencing.Small RNAs direct the RNAi and chromatin modification machinery to the cognate nucleic acids to regulate gene expression and epigenetic alterations.Notably,these acquired small RNAs and epigenetic changes persist and are transmitted from parents to offspring for multiple generations.Thus,RNAi is a vital determinant of the inheritance of gene silencing and acts as a driving force of evolution.
文摘Global prophylactic vaccination programmes have helped to curb new hepatitis B virus(HBV)infections.However,it is estimated that nearly 300 million people are chronically infected and have a high risk of developing hepatocellular carcinoma.As such,HBV remains a serious health priority and the development of novel curative therapeutics is urgently needed.Chronic HBV infection has been attributed to the persistence of the covalently closed circular DNA(cccDNA)which establishes itself as a minichromosome in the nucleus of hepatocytes.As the viral transcription intermediate,the cccDNA is responsible for producing new virions and perpetuating infection.HBV is dependent on various host factors for cccDNA formation and the minichromosome is amenable to epigenetic modifications.Two HBV proteins,X(HBx)and core(HBc)promote viral replication by modulating the cccDNA epigenome and regulating host cell responses.This includes viral and host gene expression,chromatin remodeling,DNA methylation,the antiviral immune response,apoptosis,and ubiquitination.Elimination of the cccDNA minichromosome would result in a sterilizing cure;however,this may be difficult to achieve.Epigenetic therapies could permanently silence the cccDNA minichromosome and promote a functional cure.This review explores the cccDNA epigenome,how host and viral factors influence transcription,and the recent epigenetic therapies and epigenome engineering approaches that have been described.
