Live cell CRISPR-imaging in plants reveals dynamic telomere movements
Mette, Michael F.
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AbstractElucidating the spatio-temporal organization of the genome inside the nucleus is imperative to understand the regulation of genes and non-coding sequences during development and environmental changes. Emerging techniques of chromatin imaging promise to bridge the long-standing gap between sequencing studies which reveal genomic information and imaging studies that provide spatial and temporal information of defined genomic regions. Here, we demonstrate such an imaging technique based on two orthologues of the bacterial CRISPR-Cas9 system. By fusing eGFP/mRuby2 to the catalytically inactive version of Streptococcus pyogenes and Staphylococcus aureus Cas9, we show robust visualization of telomere repeats in live leaf cells of Nicotiana benthamiana. By tracking the dynamics of telomeres visualized by CRISPR-dCas9, we reveal dynamic telomere movements of up to 2 μm within 30 minutes during interphase. Furthermore, we show that CRISPR-dCas9 can be combined with fluorescence-labelled proteins to visualize DNA-protein interactions in vivo. By simultaneously using two dCas9 orthologues, we pave the way for imaging of multiple genomic loci in live plants cells. CRISPR-imaging bears the potential to significantly improve our understanding of the dynamics of chromosomes in live plant cells.
CitationDreissig S, Schiml S, Schindele P, Weiss O, Rutten T, et al. (2017) Live cell CRISPR-imaging in plants reveals dynamic telomere movements. The Plant Journal. Available: http://dx.doi.org/10.1111/tpj.13601.
SponsorsWe would like to acknowledge Uwe Sonnewald (Friedrich-Alexander University Erlangen-Nuremberg, Germany) for providing us with pUL50-GFP. We are also thankful to Martina Dvořáčková and Jiří Fajkus (Institute of Biophysics, Brno, Czech Republic) for providing us with TRB1-GFP. This work was supported by the DFG (Grant DFG HO1779/22-1, HO1779/28-1) to A.H.) and the European Research Council (Advanced Grant COMREC 26852 to H.P.) No conflict of interest exists.
JournalThe Plant Journal