The chromosome protection mechanism under normal and replication stress environments

Dysregulation of DNA replication and consequent chromosome segregation are strongly associated with birth defects, ageing and cancer. While chromosome segregation is crucial for genome preservation, it can also result in chromosome instability, particularly when DNA replication is incomplete. Cells growing under replication-stress (RS) conditions therefore face increased risk of chromosome mis-segregation because of elevated levels of late replication intermediates (LRIs) that escape into mitosis, persistently interlinking sister chromatids and compromising their disjunction. It remains elusive how cells tolerate these aberrant conditions and suppress chromatin breakage and/or imbalanced chromatin transmission. We and others previously identified highly spatially and temporally organised LRI-associated and ultrafine-DNA bridge (UFB)-binding protein complexes that associate specifically with replication intermediate structures during mitotic progression. In this proposal, we will employ recently developed 3D Expansion STED (3D ExSTED) super-resolution nanoscopy and genome editing technologies to address the roles of, and regulatory mechanisms governing, these mitotic complexes. We aim to understand how they function in the protection and segregation of chromosomes and the pathological consequences of their deficiency. Using ExSTED, we also aim to reveal DNA folding of mitotic chromosomes in human cells. Our goal is to understand how cells maintain stable chromosome structures under normal and stressed conditions.