Triggering p53 after cytokinesis failure

Cells that fail to divide during cytokinesis often arrest in the next G1 phase by a mysterious mechanism that depends upon p53. What triggers this arrest is unclear. New studies, including a report in this issue (Uetake and Sluder, 2004) suggest that this arrest does not occur because cells are polyploid, are binucleate, have multiple centrosome, or have failed cytokinesis, making this phenomenon even more puzzling

Cell Division: Righting the Check

SummaryStudies in fission and budding yeast have continuously led the way for analyzing pathways of cell division. Two elegant studies, one from each yeast species, are opening the gates to study one of the final steps of mitosis — silencing the spindle checkpoint

The N-terminal coiled-coil of Ndel1 is a regulated scaffold that recruits LIS1 to dynein

Binding of the N terminus of Ndel1 to dynein facilitates microtubule self-organization in Ran-induced asters

Protein Architecture of the Human Kinetochore Microtubule Attachment Site

Centromeric chromatin – spindle microtubule interactions mediated by kinetochores drive chromosome segregation. We have developed a two-color fluorescence light microscopy method that measures average label separation, Delta, at < 5 nm accuracy — to elucidate the protein architecture of human metaphase kinetochores. Delta analysis, when correlated with tension states of spindle-attached sister kinetochore pairs, provided information on mechanical properties of protein linkages within kinetochores. Treatment with taxol—which suppresses microtubule dynamics, eliminates tension at kinetochores, and activates the spindle checkpoint—resulted in specific large-scale changes in kinetochore architecture. Cumulatively, Delta analysis revealed compliant linkages close to the centromeric chromatin, suggests a model for how the KMN (KNL1/Mis12 complex/Ndc80 complex) network provides microtubule attachment and generates pulling forces from depolymerization, and reveals architectural changes induced by taxol treatment. The methods described here should also be applicable to other intermediate-scale biological machines in cells

The inner centromere is a biomolecular condensate scaffolded by the chromosomal passenger complex.

The inner centromere is a region on every mitotic chromosome that enables specific biochemical reactions that underlie properties, such as the maintenance of cohesion, the regulation of kinetochores and the assembly of specialized chromatin, that can resist microtubule pulling forces. The chromosomal passenger complex (CPC) is abundantly localized to the inner centromeres and it is unclear whether it is involved in non-kinase activities that contribute to the generation of these unique chromatin properties. We find that the borealin subunit of the CPC drives phase separation of the CPC in vitro at concentrations that are below those found on the inner centromere. We also provide strong evidence that the CPC exists in a phase-separated state at the inner centromere. CPC phase separation is required for its inner-centromere localization and function during mitosis. We suggest that the CPC combines phase separation, kinase and histone code-reading activities to enable the formation of a chromatin body with unique biochemical activities at the inner centromere

Integrative single-cell meta-analysis reveals disease-relevant vascular cell states and markers in human atherosclerosis

Coronary artery disease (CAD) is characterized by atherosclerotic plaque formation in the arterial wall. CAD progression involves complex interactions and phenotypic plasticity among vascular and immune cell lineages. Single-cell RNA-seq (scRNA-seq) studies have highlighted lineage-specific transcriptomic signatures, but human cell phenotypes remain controversial. Here, we perform an integrated meta-analysis of 22 scRNA-seq libraries to generate a comprehensive map of human atherosclerosis with 118,578 cells. Besides characterizing granular cell-type diversity and communication, we leverage this atlas to provide insights into smooth muscle cell (SMC) modulation. We integrate genome-wide association study data and uncover a critical role for modulated SMC phenotypes in CAD, myocardial infarction, and coronary calcification. Finally, we identify fibromyocyte/fibrochondrogenic SMC markers (LTBP1 and CRTAC1) as proxies of atherosclerosis progression and validate these through omics and spatial imaging analyses. Altogether, we create a unified atlas of human atherosclerosis informing cell state-specific mechanistic and translational studies of cardiovascular diseases.</p

Phosphorylation Regulates SIRT1 Function

BACKGROUND: SIR2 is an NAD(+)-dependent deacetylase [1]-[3] implicated in the regulation of lifespan in species as diverse as yeast [4], worms [5], and flies [6]. We previously reported that the level of SIRT1, the mammalian homologue of SIR2 [7], [8], is coupled to the level of mitotic activity in cells both in vitro and in vivo[9]. Cells from long-lived mice maintained SIRT1 levels of young mice in tissues that undergo continuous cell replacement by proliferating stem cells. Changes in SIRT1 protein level were not associated with changes in mRNA level, suggesting that SIRT1 could be regulated post-transcriptionally. However, other than a recent report on sumoylation [10] and identification of SIRT1 as a nuclear phospho-protein by mass spectrometry [11], post-translational modifications of this important protein have not been reported. METHODOLOGY/PRINCIPAL FINDINGS: We identified 13 residues in SIRT1 that are phosphorylated in vivo using mass spectrometry. Dephosphorylation by phosphatases in vitro resulted in decreased NAD(+)-dependent deacetylase activity. We identified cyclinB/Cdk1 as a cell cycle-dependent kinase that forms a complex with and phosphorylates SIRT1. Mutation of two residues phosphorylated by Cyclin B/Cdk1 (threonine 530 and serine 540) disturbs normal cell cycle progression and fails to rescue proliferation defects in SIRT1-deficient cells [12], [13]. CONCLUSIONS/SIGNIFICANCE: Pharmacological manipulation of SIRT1 activity is currently being tested as a means of extending lifespan in mammals. Treatment of obese mice with resveratrol, a pharmacological activator of SIRT1, modestly but significantly improved longevity and, perhaps more importantly, offered some protection against the development of type 2 diabetes mellitus and metabolic syndrome [14]-[16]. Understanding the endogenous mechanisms that regulate the level and activity of SIRT1, therefore, has obvious relevance to human health and disease. Our results identify phosphorylation by cell cycle dependent kinases as a major mechanism controlling the level and function of this sirtuin and complement recent reports of factors that inhibit [17], [18] and activate [19] SIRT1 by protein-protein interactions

Spindle Poisons and Cell Fate: A Tale of Two Pathways

Spindle poisons, such as paclitaxel and vinblastine, exert their potent anti-neoplastic effects through activation of the spindle assembly checkpoint, thereby arresting cells in mitosis. Unfortunately, only certain cancers are susceptible to these drugs, and many patients fail to respond to treatment. We review the pathways that are triggered by spindle poisons and highlight recent studies that describe the great variability of tumor cells in responding to these drugs. We also describe the recent identification of an apoptotic pathway that is activated by prolonged mitotic arrest. Emerging from these studies is not only a greater understanding of how these classic antimitotic agents bring about cell death, but also a wealth of potential new targets if anticancer therapeutics