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Existing DNA stains for live cell microscopy are either toxic, require illumination with blue light, or are not compatible with super-resolution microscopy. Here the authors develop SiRHoechst, a non-toxic far-red DNA stain that is compatible with super-resolution microscopy.

Daniel Gerlich discusses how a study by the Hyman laboratory introduced the theory of liquid phase separation to cell biology and its implications for the understanding of cell organization and function.

SDS22 is a poorly characterized regulator of PP1. Here, the authors show that SDS22 prevents the aggregation of nascent PP1 and coordinates its stepwise incorporation into functional holoenzymes.

During cell division, chromosomes are maintained as individual units; this process is shown to be mediated by the cell proliferation marker Ki-67, which has biophysical properties similar to those of surfactants.

Incorporation of time information into the annotation of distinct biological states in automated fluorescence time-lapse live-cell imaging of complex cellular dynamics reduces both classification noise and confusion between cell states with similar morphology. A computational framework for achieving this is implemented in the open-source software package CellCognition.

The spindle assembly checkpoint (SAC) arrests cells in metaphase until all chromosomes are attached to the spindle. Dick and Gerlich used laser microsurgery to detach individual chromosomes, revealing that the SAC does not have a switch-like response — instead, SAC strength depends on the number of unattached chromosomes.

Histone deacetylation at the onset of mitosis induces a chromatin-intrinsic phase transition that endows chromosomes with the physical characteristics necessary for their precise movement during cell division.

Modified chromosome conformation capture (Hi-C) technology is used to characterize the interactions between sister chromatids, despite their identical DNA sequences.

Chromosomes can exist outside the nucleus in rupture-prone structures called micronuclei. It emerges that micronuclei are fragile because their outer layer lacks some nuclear-envelope components.

The surfactant-like protein Ki-67 mediates the clustering of chromosomes during mitotic exit, which displaces large cytoplasmic molecules from the future nuclear space and thus enables the separation of cytoplasmic and nuclear components before the nuclear envelope reforms.

The authors present a bioinformatic method for the accurate unsupervised classification of time-lapse images. This method should enable reproducible and unbiased annotation of large-scale image data sets.

The regulation of mitotic exit requires the rapid reversal of mitotic phosphorylation on a broad range of substrates. This requires not only inactivation of mitotic kinases but also activation of protein phosphatases, which work in regulatory networks to ensure that an interphase cell is correctly established.

Mierzwa et al. show that ESCRT-III subunit turnover at the cell midbody is driven by Vps4. Rapid turnover sustains the net growth of ESCRT-III assemblies in the presence of inhibitory Vps2 and Vps24 subunits.

Cytokinesis is the process by which mitotic cells physically split in two following chromosome segregation. Dividing animal cells first ingress a cytokinetic furrow and then separate the plasma membrane by abscission. The general cytological events and several conserved molecular factors involved in cytokinesis have been known for many years. However, recent progress in microscopy, chemical genetics, biochemical reconstitution and biophysical methodology has tremendously increased our understanding of the underlying molecular mechanisms. We discuss how recent insights have led to refined models of the distinct steps of animal cell cytokinesis, including anaphase spindle reorganization, division plane specification, actomyosin ring assembly and contraction, and abscission. We highlight how molecular signalling pathways coordinate the individual events to ensure faithful partitioning of the genome to emerging daughter cells.

DNA damage arising from replication stress is well studied, but the effect of mitotic errors on genome integrity is less understood. Here the authors knock down 47 mitotic regulators and record how they impact on DNA breakage events, providing a resource for future studies on the relation between cell division and genome integrity.

High-throughput microscopy combined with gene silencing by RNA interference is a powerful method for studying gene function. Here, a genome-wide method is presented for phenotypic screening of each of the ∼21,000 human protein-coding genes, using two-day imaging of dividing cells with fluorescently labelled chromosomes. The method enabled the identification of hundreds of genes involved in biological functions such as cell division, migration and survival.

Cells are protected from cell division errors by a pathway that detects mitotic catenation. Here, Brownlow et al.show that protein kinase Cε functions in this pathway to drive decatenation, while delaying silencing of the spindle assembly checkpoint to allow time for catenation resolution.

An RNAi screen of protein phosphatases identifies PP2A–B55α as the main mitotic exit phosphatase in human cells. After mitosis, PP2A–B55α depletion delays the reassembly of the nuclear envelope and Golgi and the decondensation of chromatin.

This protocol combines in situ chromosome conformation capture with labeling of nascent DNA with the synthetic nucleoside 4-thio-thymidine to generate genome-wide contact probability maps within and across sister chromatids in mammalian cells.

Far-red fluorogenic probes for live-cell imaging of either actin or tubulin are described and used for super-resolution microscopy of various structures in a variety of cell types.

To develop a bacterial secretion system for cytosolic delivery of therapeutic macromolecules, Chabloz et al. improve an “effectorless” Salmonella strain and combine it with a plasmid modified to boost the secretion of proteins of interest. With this system, they demonstrate efficient translocation of functional DARPins and monobodies into the cytosol of different eukaryotic cells lines and successfully block the paradigmatic RAS pathway.