The conformation of replicated human chromosomes
The three-dimensional conformation of chromatids has tremendous impact on a variety of different aspects of cell biology such as gene expression, chromosome segregation and DNA repair. High-throughput chromosome conformation capture (Hi-C) has enabled researchers to examine this conformation in great detail and has lead to the discovery of numerous principles of chromosome topology. However, Hi-C is currently not able to distinguish replicated chromosomes, thus precluding investigators from
... estigators from examining key aspects of genome function. These include how DNA repair coordinates copying information from sister chromatids and how sister chromatid resolution during mitosis is coordinated with sister chromatid cohesion. In order to resolve this methodological conundrum, I used 4-thio-thymidine, a novel nucleotide analog that is detectable by sequencing, to mark sister chromatids differentially. This enabled me to develop a Hi-C variant, which I name sister chromatid sensitive Hi-C (scsHi-C), that is capable of distinguishing sister chromatids. I then used this method to generate the first sister chromatid resolved 3D-genome maps, revealing that trans-sister interactions exhibit remarkable heterogeneity. I further show that sister chromatids are held together at the boundaries of topologically associating domains and that trans-sister interactions correlate with epigenetic state. Finally, I use perturbations of the cohesin complex to show that two classes of cohesin organize sister chromatids differentially: Cohesive cohesin holds sister chromatids together, whereas loop extruding cohesin separates them on a small genomic scale. In conclusion, scsHi-C is a novel technology that allows sister chromatid specific 3D-structure examinations that I showcased by examining the influence of the cohesin complex on this configuration.