TY - JOUR A1 - Yan, Wenhao A1 - Chen, Dijun A1 - Schumacher, Julia A1 - Durantini, Diego A1 - Engelhorn, Julia A1 - Chen, Ming A1 - Carles, Cristel C. A1 - Kaufmann, Kerstin T1 - Dynamic control of enhancer activity drives stage-specific gene expression during flower morphogenesis JF - Nature Communications N2 - Enhancers are critical for developmental stage-specific gene expression, but their dynamic regulation in plants remains poorly understood. Here we compare genome-wide localization of H3K27ac, chromatin accessibility and transcriptomic changes during flower development in Arabidopsis. H3K27ac prevalently marks promoter-proximal regions, suggesting that H3K27ac is not a hallmark for enhancers in Arabidopsis. We provide computational and experimental evidence to confirm that distal DNase. hypersensitive sites are predictive of enhancers. The predicted enhancers are highly stage-specific across flower development, significantly associated with SNPs for flowering-related phenotypes, and conserved across crucifer species. Through the integration of genome-wide transcription factor (TF) binding datasets, we find that floral master regulators and stage-specific TFs are largely enriched at developmentally dynamic enhancers. Finally, we show that enhancer clusters and intronic enhancers significantly associate with stage-specific gene regulation by floral master TFs. Our study provides insights into the functional flexibility of enhancers during plant development, as well as hints to annotate plant enhancers. Y1 - 2019 U6 - https://doi.org/10.1038/s41467-019-09513-2 SN - 2041-1723 VL - 10 PB - Nature Publ. Group CY - London ER - TY - JOUR A1 - Yan, Wenhao A1 - Chen, Dijun A1 - Kaufmann, Kerstin T1 - Efficient multiplex mutagenesis by RNA-guided Cas9 and its use in the characterization of regulatory elements in the AGAMOUS gene JF - Plant methods N2 - Background The efficiency of multiplex editing in plants by the RNA-guided Cas9 system is limited by efficient introduction of its components into the genome and by their activity. The possibility of introducing large fragment deletions by RNA-guided Cas9 tool provides the potential to study the function of any DNA region of interest in its ‘endogenous’ environment. Results Here, an RNA-guided Cas9 system was optimized to enable efficient multiplex editing in Arabidopsis thaliana. We demonstrate the flexibility of our system for knockout of multiple genes, and to generate heritable large-fragment deletions in the genome. As a proof of concept, the function of part of the second intron of the flower development gene AGAMOUS in Arabidopsis was studied by generating a Cas9-free mutant plant line in which part of this intron was removed from the genome. Further analysis revealed that deletion of this intron fragment results 40 % decrease of AGAMOUS gene expression without changing the splicing of the gene which indicates that this regulatory region functions as an activator of AGAMOUS gene expression. Conclusions Our modified RNA-guided Cas9 system offers a versatile tool for the functional dissection of coding and non-coding DNA sequences in plants. KW - RNA-guided Cas9 KW - Multiplex mutagenesis KW - Large fragment deletion KW - Germline transmission Y1 - 2016 U6 - https://doi.org/10.1186/s13007-016-0125-7 SN - 1746-4811 VL - 12 SP - 1 EP - 9 PB - BioMed Central CY - London ER - TY - JOUR A1 - Yan, Wenhao A1 - Chen, Dijun A1 - Kaufmann, Kerstin T1 - Efficient multiplex mutagenesis by RNA-guided Cas9 and its use in the characterization of regulatory elements in the AGAMOUS gene JF - Plant Methods N2 - Results: Here, an RNA-guided Cas9 system was optimized to enable efficient multiplex editing in Arabidopsis thaliana. We demonstrate the flexibility of our system for knockout of multiple genes, and to generate heritable large-fragment deletions in the genome. As a proof of concept, the function of part of the second intron of the flower development gene AGAMOUS in Arabidopsis was studied by generating a Cas9-free mutant plant line in which part of this intron was removed from the genome. Further analysis revealed that deletion of this intron fragment results 40 % decrease of AGAMOUS gene expression without changing the splicing of the gene which indicates that this regulatory region functions as an activator of AGAMOUS gene expression. Conclusions: Our modified RNA-guided Cas9 system offers a versatile tool for the functional dissection of coding and non-coding DNA sequences in plants. KW - RNA-guided Cas9 KW - Multiplex mutagenesis KW - Large fragment deletion KW - Germline transmission Y1 - 2016 U6 - https://doi.org/10.1186/s13007-016-0125-7 SN - 1746-4811 VL - 12 SP - 2381 EP - 2389 PB - BioMed Central CY - London ER - TY - JOUR A1 - Yan, Wenhao A1 - Chen, Dijun A1 - Kaufmann, Kerstin T1 - Molecular mechanisms of floral organ specification by MADS domain proteins JF - Current opinion in plant biology N2 - Flower development is a model system to understand organ specification in plants. The identities of different types of floral organs are specified by homeotic MADS transcription factors that interact in a combinatorial fashion. Systematic identification of DNA-binding sites and target genes of these key regulators show that they have shared and unique sets of target genes. DNA binding by MADS proteins is not based on ‘simple’ recognition of a specific DNA sequence, but depends on DNA structure and combinatorial interactions. Homeotic MADS proteins regulate gene expression via alternative mechanisms, one of which may be to modulate chromatin structure and accessibility in their target gene promoters. Y1 - 2016 U6 - https://doi.org/10.1016/j.pbi.2015.12.004 SN - 1369-5266 SN - 1879-0356 VL - 29 SP - 154 EP - 162 PB - Elsevier CY - London ER - TY - GEN A1 - Yan, Wenhao A1 - Chen, Dijun A1 - Kaufmann, Kerstin T1 - Efficient multiplex mutagenesis by RNA‑guided Cas9 and its use in the characterization of regulatory elements in the AGAMOUS gene N2 - Background: The efficiency of multiplex editing in plants by the RNA-guided Cas9 system is limited by efficient introduction of its components into the genome and by their activity. The possibility of introducing large fragment deletions by RNA-guided Cas9 tool provides the potential to study the function of any DNA region of interest in its ‘endogenous’ environment. Results: Here, an RNA-guided Cas9 system was optimized to enable efficient multiplex editing in Arabidopsis thaliana. We demonstrate the flexibility of our system for knockout of multiple genes, and to generate heritable largefragment deletions in the genome. As a proof of concept, the function of part of the second intron of the flower development gene AGAMOUS in Arabidopsis was studied by generating a Cas9-free mutant plant line in which part of this intron was removed from the genome. Further analysis revealed that deletion of this intron fragment results 40 % decrease of AGAMOUS gene expression without changing the splicing of the gene which indicates that this regulatory region functions as an activator of AGAMOUS gene expression. Conclusions: Our modified RNA-guided Cas9 system offers a versatile tool for the functional dissection of coding and non-coding DNA sequences in plants. T3 - Zweitveröffentlichungen der Universität Potsdam : Mathematisch-Naturwissenschaftliche Reihe - 224 KW - RNA-guided Cas9 KW - Multiplex mutagenesis KW - Large fragment deletion KW - Germline transmission Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:517-opus4-90895 ER -