Figure 4.

Chromosome compartmentalization dynamics and cell fate decision during mitosis. A, LLPS-driven central spindle assembly by membraneless organelles requires CENP-E and BubR1 kinase. Representative phenotypes of the metaphase-anaphase transition in HeLa cells treated with bubristatin-1 (BRT-1). HeLa cells expressing GFP-tubulin and mCherry-H2B were synchronized at metaphase with MG132. Once released from metaphase arrest, cells were treated with BRT-1 or DMSO followed by real-time imaging with deconvolution microscopy. A current interest is to visualize how CENP-E-BubR1 coacervates regulate BubR1 kinase activity in the central spindle. Scale bar, 10 μm. Adapted from Ref. 74. B, LLPS-driven centromere kinase Mps1 (monopolar spindle 1) coacervates are essential for stable kinetochore localization in mitosis. HeLa cells expressing LAP-Mps1 were exposed to DMSO (bottom) or reversine (an Mps1 inhibitor; top) in the presence of MG132 and nocodazole for synchronization, before being fixed and counterstained for anti-centromere antibody (ACA; shown as red in merged images) and DNA (blue). It was apparent that Mps1 kinase coacervates are stable in the presence of reversine, an inhibitor for preventing Mps1 from utilizing ATP, indicating that Mps1 autophosphorylation liberates Mps1 coacervates from kinetochores. A current interest is to determine whether coacervate dissolution is regulated by Mps1 or its downstream effectors. Scale bar, 10 μm. Adapted from Refs. 89 and 110.