Visualizing chromosomal rearrangements caused by replication fork stalling in a single cell

Aberrant chromosome structures can promote tumors in the early stages of carcinogenesis and lead to tumor cells becoming resistant to chemotherapy, for example by changing in drug metabolism. Dicentric (containing two centromeres) and acentric (containing no centromeres) chromosomes are two abnormal chromosome structures that consider as precursors of a variety of gross chromosomal rearrangements (GCRs) generated by subsequent recombination events [1-7]. However, the mechanism of the dicentric and acentric palindromic chromosome formation and their subsequent metabolism is difficult to directly visualise. The previous results from our lab shows that replication forks stalled at a specific replication termination sequence (RTS1) can result in the formation of the dicentric and acentric palindromic chromosomes in the fission yeast Schizosaccharomyces pombe [48-52]. However, the formation of acentric and dicentric chromosomes results in a significant visability loss, due to instability and miss-segregation of the chromosomes in the yeast cells. Thus, their fate is difficult or impossible to follow. To resolve this problem, a non-essential mini-chromosome (Ch16) was developed as a novel model system in this project. The behaviour of rearranged chromosome in vivo and their subsequent fate have been visualised by integrating the lac operator (lacO) and tetracycline operator (tetO) arrays with auxotropic makers, adjacent to the RTSI locus on Ch16. The results reveal imbalanced segregation of a dicentric chromosome and subsequently undergoes a breakage event. An acentric chromosome appears to be decoupled or lost rapidly from the nucleus.