biostudies-literatureWang MNational Nonprofit Institute Research Grant of the Chinese Academy of ForestryNational Natural Science Foundation of Chinathe National Natural Science Foundation of Chinathe National Nonprofit Institute Research Grant of the Chinese Academy of Forestry10492https://www.ebi.ac.uk/biostudies/studies/S-EPMC9504898biostudies-literatureUnknown23(18)CRISPR/dCas9 is an important DNA modification tool in which a disarmed Cas9 protein with no nuclease activity is fused with a specific DNA modifying enzyme. A previous study reported that overexpression of the TET1 catalytic domain (TET1cd) reduces genome-wide methylation in Arabidopsis. A spontaneous naturally occurring methylation region (NMR19-4) was identified in the promoter region of the <i>PPH</i> (<i>Pheophytin Pheophorbide Hydrolase</i>) gene, which encodes an enzyme that can degrade chlorophyll and accelerate leaf senescence. The methylation status of NMR19-4 is associated with <i>PPH</i> expression and leaf senescence in Arabidopsis natural accessions. In this study, we show that the CRISPR/dCas9-TET1cd system can be used to target the methylation of hypermethylated NMR19-4 region to reduce the level of methylation, thereby increasing the expression of <i>PPH</i> and accelerating leaf senescence. Furthermore, hybridization between transgenic demethylated plants and hypermethylated ecotypes showed that the demethylation status of edited NMR19-4, along with the enhanced <i>PPH</i> expression and accelerated leaf senescence, showed Mendelian inheritance in F1 and F2 progeny, indicating that spontaneous epialleles are stably transmitted trans-generationally after demethylation editing. Our results provide a rational approach for future editing of spontaneously mutated epialleles and provide insights into the epigenetic mechanisms that control plant leaf senescence.International journal of molecular sciencesTransgenerationally Transmitted DNA Demethylation of a Spontaneous Epialleles Using CRISPR/dCas9-TET1cd Targeted Epigenetic Editing in Arabidopsis.PMC950489831871220CAFYBB2019ZY003Wang MCao LZhang TXie LZhou WHe LWang YChen BZhang QWang LZhang PfalseTransgenerationally Transmitted DNA Demethylation of a Spontaneous Epialleles Using CRISPR/dCas9-TET1cd Targeted Epigenetic Editing in Arabidopsis.CRISPR/dCas9 is an important DNA modification tool in which a disarmed Cas9 protein with no nuclease activity is fused with a specific DNA modifying enzyme. A previous study reported that overexpression of the TET1 catalytic domain (TET1cd) reduces genome-wide methylation in Arabidopsis. A spontaneous naturally occurring methylation region (NMR19-4) was identified in the promoter region of the <i>PPH</i> (<i>Pheophytin Pheophorbide Hydrolase</i>) gene, which encodes an enzyme that can degrade chlorophyll and accelerate leaf senescence. The methylation status of NMR19-4 is associated with <i>PPH</i> expression and leaf senescence in Arabidopsis natural accessions. In this study, we show that the CRISPR/dCas9-TET1cd system can be used to target the methylation of hypermethylated NMR19-4 region to reduce the level of methylation, thereby increasing the expression of <i>PPH</i> and accelerating leaf senescence. Furthermore, hybridization between transgenic demethylated plants and hypermethylated ecotypes showed that the demethylation status of edited NMR19-4, along with the enhanced <i>PPH</i> expression and accelerated leaf senescence, showed Mendelian inheritance in F1 and F2 progeny, indicating that spontaneous epialleles are stably transmitted trans-generationally after demethylation editing. Our results provide a rational approach for future editing of spontaneously mutated epialleles and provide insights into the epigenetic mechanisms that control plant leaf senescence.2022-01-01T00:00:00Z2022 Sep2026-04-08T11:50:51.73Z2025-02-19T00:44:13.444ZS-EPMC95048983614240710.3390/ijms231810492