Deconstructing the molecular basis of condensin-mediated chromatin folding
Abstract: The view of genes and regulatory elements along a straight line has been found lacking. We now know that the genome functions as a three-dimensional chromatin polymer, driven by a complex collection of chromosome interactions at multiple scales, from small chromatin loops to the segregation of whole chromosomes into distinct territories. And yet, the mechanisms that control these interactions and drive the spatial organization of the genome remain unknown. To address this gap, we have recently developed two technologies that use fluorescent in situ hybridization (FISH) to interrogate chromosome interactions at single-cell resolution. The first is a technology for high-throughput FISH (Hi-FISH), and the other, called Oligopaints, is a new type of probe that reduces the cost and increases the resolution of FISH. Using these tools, our work has provided new insights into how chromosomes find and influence each other in the nucleus. In particular, we and others have isolated the condensin II complex as a major organizing factor that can drive chromatin compaction and chromosome separation during interphase. We hypothesize that the intra-chromosomal functions of compaction and chromatin looping may be a mechanism by which long-range and inter-chromosomal interactions are inhibited. We propose to build on this work and determine how condensin II drives the folding of interphase chromosomes using our Oligopaints technology to generate low-cost chromosome paints for systematic analyses of chromosome size, morphology, and position. The studies proposed here will advance our fundamental understanding of condensin biology, providing a new avenue to study how chromosome positioning is established and inherited, and how dysfunctional organization contributes to genome instability.