1,6-hexanediol rapidly immobilizes and condenses chromatin in living human cells

Abstract

Alcohol, 1,6-hexanediol, is widely used for melting liquid droplets formed by liquid0–liquid phase separation; however, it can also seriously affect cells by immobilizing and condensing chromatin. Liquid droplets formed inside the cell by liquid–liquid phase separation maintain membrane-less condensates/bodies (or compartments). These droplets are important for concentrating certain molecules and facilitating spatiotemporal regulation of cellular functions. 1,6-hexanediol (1,6-HD), an aliphatic alcohol, inhibits weak hydrophobic protein–protein/protein-RNA interactions required for the droplet formation (droplet melting activity) and is used here to elucidate the formation process of cytoplasmic/nuclear condensates/bodies. However, the effect of 1,6-HD on chromatin in living cells remains unclear. We found that 1,6-HD drastically suppresses chromatin motion and hyper-condenses chromatin in human cells by using live-cell single-nucleosome imaging, which detects changes in the state of chromatin. These effects were enhanced in a dose-dependent manner. Chromatin was “frozen” by 5%, or higher, concentrations of 1,6-HD. 1,6-HD greatly facilitated cation-dependent chromatin condensation in vitro. This 1,6-HD action is distinct from its melting activity of liquid droplets. Alcohols, such as 1,6-HD, appear to remove water molecules around chromatin and locally condense chromatin. Therefore, liquid droplet results obtained using 1,6-HD should be carefully interpreted or reconsidered when these droplets are associated with chromatin.