Heterochromatin provides an epigenetically controlled method of silencing genes and maintaining chromosome structure. In the fission yeast Schizosaccharomyces pombe , the HP1 homolog Swi6 plays an important role in heterochromatin establishment and maintenance. Swi6 has two conserved domains that have been extensively studied, but its two intrinsically disordered regions (IDRs) are poorly understood. The IDRs are difficult to study because Swi6 uses them to control short-term interactions with several different binding partners that combine to form biomolecular condensates, rendering in vitro techniques incapable of building a complete picture of how Swi6 uses its IDRs. Here, we study the role of the Swi6 IDRs in living cells using single-particle tracking PALM (sptPALM) of Swi6-PAmCherry. We have identified Swi6 IDR variants that cause a change in heterochromatin maintenance, and we have used our single-molecule measurements to compare the behavior of each Swi6 IDR variant. Furthermore, we have quantified the velocity, heterogeneity, and anisotropy of the Swi6 trajectories to determine differences in heterochromatin binding by the different Swi6 IDR variants. Our work highlights the importance of IDRs, which are often overlooked, and demonstrates how single-molecule microscopy can measure the behavior of proteins in live cells.
Steen et al. (Sun,) studied this question.