Chemically Induced Chromosomal Interaction (CICI) method to study chromosome dynamics and its biological roles

Abstract

Numerous intra- and inter-chromosomal contacts have been mapped in eukaryotic genomes, but it remains challenging to link these 3D structures to their regulatory functions. To establish the causal relationships between chromosome conformation and genome functions, we develop a method, Chemically Induced Chromosomal Interaction (CICI), to selectively perturb the chromosome conformation at targeted loci. Using this method, long-distance chromosomal interactions can be induced dynamically between intra- or inter-chromosomal loci pairs, including the ones with very low Hi-C contact frequencies. Measurement of CICI formation time allows us to probe chromosome encounter dynamics between different loci pairs across the cell cycle. We also conduct two functional tests of CICI. We perturb the chromosome conformation near a DNA double-strand break and observe altered donor preference in homologous recombination; we force interactions between early and late-firing DNA replication origins and find no significant changes in replication timing. These results suggest that chromosome conformation plays a deterministic role in homology-directed DNA repair, but not in the establishment of replication timing. Overall, our study demonstrates that CICI is a powerful tool to study chromosome dynamics and 3D genome function.

DNA Methylation-Based High-Resolution Mapping of Long-Distance Chromosomal Interactions in Nucleosome-Depleted Regions

“…The two mating types of budding yeast, a and α, are controlled by sequences in the mating locus (MAT) on chromosome III (chrIII). During each mother cell cycle, HO endonuclease is expressed and generates a double strand break (DSB) in MAT, which is repaired by a homologous sequence, HMLα or To further corroborate our findings, we forced the cells to switch mating type by expressing the HO endonuclease controlled by the GAL1 promoter 40,41 . Following the procedure in…”

“…To further corroborate our findings, we forced the cells to switch mating type by expressing the HO endonuclease controlled by the GAL1 promoter 40,41 . Following the procedure in Fig S5A , we generated a pulse of HO expression that led to the mating-type switch in ∼47% of the cells ( Fig S5B & C ).…”

“…This fusion gene is driven by an engineered core GAL1 promoter that binds to LexA-ER-VP16, which can be transcriptionally induced by β-estradiol 30,54 . MTAC targeted strains were constructed by integrating a plasmid containing 256X LacO repeats derived from a previous study 41 into the corresponding VP locus into the control strain.…”

“…The experiment was performed as previously described 41 . Single colonies were grown to log-phase in 50 mL synthetic media with raffinose.…”

JACKIE: Fast Enumeration of Genome-Wide Single- and Multicopy CRISPR Target Sites and Their Off-Target Numbers

“…Furthermore, no design software is available for genome imaging or loop engineering experiments requiring clustered repetitive sequences. 4 , 5 , 7 , 10 , 12–14 , 17 …”