Quantifying cell-cycle-dependent chromatin dynamics during interphase by live 3D tracking

The study of cell cycle progression and regulation is important to our understanding of fundamental biophysics, aging, and disease mechanisms. Local chromatin movements are generally considered to be constrained and relatively consistent during all interphase stages, although recent advances in our understanding of genome organization challenge this claim. Here, we use high spatiotemporal resolution, 4D (x, y, zand time) localization microscopyby point-spread-function (PSF) engineering and deep learning-based image analysis, for live imaging of mouse embryonic fibroblast (MEF3T3) and MEF 3T3 double Lamin A Knockout (LmnaKO) cell lines, to characterizetelomere diffusion during the interphase. We detected varying constraint levels imposed on chromatin, which are prominently decreased duringG0/G1. Our 4D measurements of telomere diffusion offer an effectivemethod to investigate chromatin dynamics and reveal cell-cycle-dependent motion constraints, which may be caused by various cellular processes.