4D reorganization of yeast megachromsomes unveils spatio-temporal aspects of replication firing and gene contacts in yeast

Eukaryotic genomes vary greatly in terms of size, chromosome number and genetic complexity. Here we used engineered karyotypes of budding yeast – in which 16 native chromosomes were fused into two gigantic DNA molecules - to characterize effects of chromosome length on nuclear architecture and function. We reasoned that a ~5-fold increase in chromosome length - relative to the longest native chromosome - might alter nuclear occupancy of the megachromosomes. To investigate this hypothesis, we measured the proportion of the nucleus occupied by genomic DNA in cells labeled with SYTOX Green (see methods below). We observed that megachromosomes occupy a larger fraction of nuclear volume than native chromosomes that was correlated with an increase in the size of the nucleus.