Live-Cell Imaging of Chromatin Condensation Dynamics by CRISPR

• Published in: iScience Vol. 4; pp. 216 - 235
• Main Authors: Xue, Yuan, Acar, Murat
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 29.06.2018; Elsevier
• Subjects: Chromosome Organization; Genetics; Live Imaging; Molecular Biology; Techniques in Genetics; Genetics; Techniques in Genetics; Chromosome Organization; Molecular Biology; Live Imaging
• ISSN: 2589-0042; 2589-0042
• DOI: 10.1016/j.isci.2018.06.001

The spatiotemporal organization of chromatin plays central roles in cellular function. The ribosomal DNA (rDNA) chromatin undergoes dynamic structural changes during mitosis and stress. Here, we developed a CRISPR-based imaging system and tracked the condensation dynamics of rDNA chromatin in live yeast cells under glucose starvation. We found that acute glucose starvation triggers rapid condensation of rDNA. Time-lapse microscopy revealed two stages for rDNA condensation: a “primary stage,” when relaxed rDNA chromatin forms higher order loops or rings, and a “secondary stage,” when the rDNA rings further condense into compact clusters. Twisting of rDNA rings accompanies the secondary stage. The condensin complex, but not the cohesin complex, is required for efficient rDNA condensation in response to glucose starvation. Furthermore, we found that the DNA helicase Sgs1 is essential for the survival of cells expressing rDNA-bound dCas9, suggesting a role for helicases in facilitating DNA replication at dCas9-binding sites. [Display omitted] •A CRISPR-based imaging system allows tracking of rDNA condensation in single cells•Glucose starvation triggers rDNA condensation in two prominent stages•Condensin contributes to efficient rDNA condensation caused by glucose starvation•Sgs1 helicase is required for normal rDNA replication at dCas9-binding sites Genetics; Techniques in Genetics; Molecular Biology; Chromosome Organization; Live Imaging