Histone citrullination,an important post-translational modification mediated by peptidyl arginine deiminases,is essential for many physiological processes and epigenetic regulation.However,the causal relationship betw...Histone citrullination,an important post-translational modification mediated by peptidyl arginine deiminases,is essential for many physiological processes and epigenetic regulation.However,the causal relationship between histone citrullination and specific gene regulation remains unresolved.In this study,we develop a programmable epigenetic editor by fusing the peptidyl arginine deiminase(PAD)PPAD from Porphyromonas gingivalis with d Cas9.With the assistance of g RNA,PPAD-d Cas9 can recruit PPADs to specific genomic loci,enabling direct manipulation of the epigenetic landscape and regulation of gene expression.Our citrullination editor allows for the site-specific manipulation of histone H3R2,8,17 and H3R26 at target human gene loci,resulting in the activation or suppression of different genes in a locus-specific manner.Moreover,the epigenetic effects of the citrullination editor are specific and sustained.This epigenetic editor offers an accurate and efficient tool for exploring gene regulation of histone citrullination.展开更多
Dear Editor,CRISPR-Cas9 （clustered regularly interspaced short palin- dromic repeats-CRISPR associated） systems have been harnessed for kinds of genome manipulation, including gene editing, transcription regulation,...Dear Editor,CRISPR-Cas9 （clustered regularly interspaced short palin- dromic repeats-CRISPR associated） systems have been harnessed for kinds of genome manipulation, including gene editing, transcription regulation, and chromosome loci imaging （Dominguez et al., 2016; Komor et al., 2017）. A typical engineered CRISPR-Cas9 system is composed of a Cas9 protein and a single guide RNA （sgRNA）, which could form a protein/RNA complex to recognize and cleave DNA sequence （Hsu et al., 2014; Wright et al., 2016）.展开更多
Synthetic gene activators consisting of nucleasedead Cas9(dCas9)for single-guide RNA(sgRNA)-directed promoter binding and a transcriptional activation domain(TAD)represent new tools for gene activation from endogenous...Synthetic gene activators consisting of nucleasedead Cas9(dCas9)for single-guide RNA(sgRNA)-directed promoter binding and a transcriptional activation domain(TAD)represent new tools for gene activation from endogenous genomic locus in basic and applied plant research.However,multiplex gene coactivation by d Cas9-TADs has not been demonstrated in whole plants.There is also room to optimize the performance of these tools.Here,we report that our previously developed gene activator,dCas9-TV,could simultaneously upregulate OsGW7 and OsER1 in rice by up to 3,738 fold,with one sg RNA targeting to each promoter.The gene coactivation could persist to at least the fourth generation.Astonishingly,thepolycistronictRNA-sgRNAexpression under the maize ubiquitin promoter,a Pol II promoter,could cause enormous activation of these genes by up to>40,000-fold in rice.Moreover,the yeast GCN4 coiled coil-mediated dCas9-TV dimerization appeared to be promising for enhancing gene activation.Finally,we successfully introduced a self-amplification loop for dCas9-TV expression in Arabidopsis to promote the transcriptional upregulation of AtFLS2,a previously characterized dCas9-TV-refractory gene with considerable basal expression.Collectively,this work illustrates the robustness of dCas9-TV in multigene coactivation and provides broadly useful strategies for boosting transcriptional activation efficacy of dCas9-TADs in plants.展开更多
基金funded by the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project(TSBICIP-CXRC-048)。
文摘Histone citrullination,an important post-translational modification mediated by peptidyl arginine deiminases,is essential for many physiological processes and epigenetic regulation.However,the causal relationship between histone citrullination and specific gene regulation remains unresolved.In this study,we develop a programmable epigenetic editor by fusing the peptidyl arginine deiminase(PAD)PPAD from Porphyromonas gingivalis with d Cas9.With the assistance of g RNA,PPAD-d Cas9 can recruit PPADs to specific genomic loci,enabling direct manipulation of the epigenetic landscape and regulation of gene expression.Our citrullination editor allows for the site-specific manipulation of histone H3R2,8,17 and H3R26 at target human gene loci,resulting in the activation or suppression of different genes in a locus-specific manner.Moreover,the epigenetic effects of the citrullination editor are specific and sustained.This epigenetic editor offers an accurate and efficient tool for exploring gene regulation of histone citrullination.
文摘Dear Editor,CRISPR-Cas9 （clustered regularly interspaced short palin- dromic repeats-CRISPR associated） systems have been harnessed for kinds of genome manipulation, including gene editing, transcription regulation, and chromosome loci imaging （Dominguez et al., 2016; Komor et al., 2017）. A typical engineered CRISPR-Cas9 system is composed of a Cas9 protein and a single guide RNA （sgRNA）, which could form a protein/RNA complex to recognize and cleave DNA sequence （Hsu et al., 2014; Wright et al., 2016）.
基金supported by a grant from the National Transgenic Science and Technology Major Program of China(2019ZX08010003-001-009)。
文摘Synthetic gene activators consisting of nucleasedead Cas9(dCas9)for single-guide RNA(sgRNA)-directed promoter binding and a transcriptional activation domain(TAD)represent new tools for gene activation from endogenous genomic locus in basic and applied plant research.However,multiplex gene coactivation by d Cas9-TADs has not been demonstrated in whole plants.There is also room to optimize the performance of these tools.Here,we report that our previously developed gene activator,dCas9-TV,could simultaneously upregulate OsGW7 and OsER1 in rice by up to 3,738 fold,with one sg RNA targeting to each promoter.The gene coactivation could persist to at least the fourth generation.Astonishingly,thepolycistronictRNA-sgRNAexpression under the maize ubiquitin promoter,a Pol II promoter,could cause enormous activation of these genes by up to>40,000-fold in rice.Moreover,the yeast GCN4 coiled coil-mediated dCas9-TV dimerization appeared to be promising for enhancing gene activation.Finally,we successfully introduced a self-amplification loop for dCas9-TV expression in Arabidopsis to promote the transcriptional upregulation of AtFLS2,a previously characterized dCas9-TV-refractory gene with considerable basal expression.Collectively,this work illustrates the robustness of dCas9-TV in multigene coactivation and provides broadly useful strategies for boosting transcriptional activation efficacy of dCas9-TADs in plants.
