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53BP1 acts at the crossroads between DNA repair and p53‐mediated stress response. With its interactors p53 and USP28, it is part of the mitotic surveillance (or mitotic stopwatch) pathway (MSP), a sensor that monitors the duration of cell division, promoting p53‐dependent cell cycle arrest when a critical time threshold is surpassed. Here, we show that Polo‐like kinase 1 (PLK1) activity is essential for the time‐dependent release of 53BP1 from kinetochores. PLK1 inhibition, which leads to 53BP1 persistence at kinetochores, prevents cytosolic 53BP1 association with p53 and results in a blunted MSP. Strikingly, the identification of CENP‐F as the kinetochore docking partner of 53BP1 enabled us to show that measurement of mitotic timing by the MSP does not take place at kinetochores, as perturbing CENP‐F‐53BP1 binding had no measurable impact on the MSP. Taken together, we propose that PLK1 supports the MSP by generating a cytosolic pool of 53BP1 and that an unknown cytosolic mechanism enables the measurement of mitotic duration. Synopsis: Following an abnormally extended mitosis, cells activate p53 via 53BP1. Surprisingly, 53BP1 kinetochore localization is dispensable for this process, whereas PLK1 activity, a negative regulator of 53BP1 localization, appears to be essential.53BP1 kinetochore localization is dispensable for the mitotic surveillance (or mitotic stopwatch) pathway (MSP).PLK1 is required for the MSP.PLK1 activity promotes 53BP1 kinetochore dynamics and 53BP1‐p53 complex formation. [ABSTRACT FROM AUTHOR]

Unscheduled increases in ploidy underlie defects in tissue function, premature aging, and malignancy. A concomitant event to polyploidization is the amplification of centrosomes, the main microtubule organization centers in animal cells. Supernumerary centrosomes are frequent in tumors, correlating with higher aggressiveness and poor prognosis. However, extra centrosomes initially also exert an onco‐protective effect by activating p53‐induced cell cycle arrest. If additional signaling events initiated by centrosomes help prevent pathology is unknown. Here, we report that extra centrosomes, arising during unscheduled polyploidization or aberrant centriole biogenesis, induce activation of NF‐κB signaling and sterile inflammation. This signaling requires the NEMO‐PIDDosome, a multi‐protein complex composed of PIDD1, RIPK1, and NEMO/IKKγ. Remarkably, the presence of supernumerary centrosomes suffices to induce a paracrine chemokine and cytokine profile, able to polarize macrophages into a pro‐inflammatory phenotype. Furthermore, extra centrosomes increase the immunogenicity of cancer cells and render them more susceptible to NK‐cell attack. Hence, the PIDDosome acts as a dual effector, able to engage not only the p53 network for cell cycle control but also NF‐κB signaling to instruct innate immunity. Synopsis: Centrosome accumulation in tumors correlates with poor prognosis but can initially also induce antitumorigenic p53 responses. Here, extra centrosomes are found to also trigger NF‐kB signaling via the NEMO‐PIDDosome, thereby increasing immunogenicity of cancer cells. Extra centrosomes can elicit autocrine and paracrine inflammatory signals.The NEMO‐PIDDosome induces NF‐kB signaling downstream of extra centrosomes.Cancer cells with extra centrosomes become vulnerable to attack by natural killer (NK) cells.Vulnerability to NK cell‐killing depends on PIDD1‐induced NF‐kB activation. [ABSTRACT FROM AUTHOR]

Background: According to the REACH (Registration, Evaluation, Authorisation and Restriction of Chemicals) restriction, tattoo and permanent make‐up (PMU) inks placed on the European Union market after January 4, 2022, shall not contain methylisothiazolinone, benzisothiazolinone (BIT), octylisothiazolinone (OIT), or other skin sensitizers in concentrations of 10 mg/kg or higher and phenoxyethanol (PE) or other eye irritants or damaging substances in concentrations of 100 mg/kg or higher. In addition, preservatives and other substances enlisted in Annex II to Cosmetic Product Regulation shall not be present in concentrations of 0.5 mg/kg or higher. Objectives: This study aims to quantify 14 preservatives in 99 tattoo and 39 PMU inks from the Italian market and presents a comparison with concentration limits set by the REACH restriction. Methods: Inks were analysed by applying validated analytical methods based on liquid chromatography techniques. Results: About 24.0%, 15.2% and 1.5% of the overall samples contained BIT, PE and OIT, respectively, at concentrations exceeding REACH concentration limits. The number of noncompliant tattoo inks (49.5%) would be significantly greater than that of the PMU inks (17.9%). Conclusions: About 40.6% of the samples would be noncompliant with the restriction for the presence of preservatives above the permitted level. Additional concentration limits will apply to skin sensitizing preservatives for proper labelling of inks under CLP (Classification, Labelling and Packaging) Regulation. [ABSTRACT FROM AUTHOR]

A study was conducted on the cloning and sequencing of the atpHAGDC operon of Rhodobacter capsulatus, containing the five genes coding for the F1 sector of the ATP synthase. The results indicate that R capsulatus grows poorly under pure fermentative conditions with fructose as the carbon source. The findings also suggest that anaerobic electron flow is associated with oxidative phosphorylation.

Centrosome amplification results into genetic instability and predisposes cells to neoplastic transformation. Supernumerary centrosomes trigger p53 stabilization dependent on the PIDDosome (a multiprotein complex composed by PIDD1, RAIDD and Caspase‐2), whose activation results in cleavage of p53's key inhibitor, MDM2. Here, we demonstrate that PIDD1 is recruited to mature centrosomes by the centriolar distal appendage protein ANKRD26. PIDDosome‐dependent Caspase‐2 activation requires not only PIDD1 centrosomal localization, but also its autoproteolysis. Following cytokinesis failure, supernumerary centrosomes form clusters, which appear to be necessary for PIDDosome activation. In addition, in the context of DNA damage, activation of the complex results from a p53‐dependent elevation of PIDD1 levels independently of centrosome amplification. We propose that PIDDosome activation can in both cases be promoted by an ANKRD26‐dependent local increase in PIDD1 concentration close to the centrosome. Collectively, these findings provide a paradigm for how centrosomes can contribute to cell fate determination by igniting a signalling cascade. SYNOPSIS: The PIDDosome is a Caspase‐2‐activating platform assembling in response to two stimuli: genotoxic stress and supernumerary centrosomes. Here, both triggers are shown to rely on centrosomal localization of PIDD1, highlighting the centrosome's capability to influence cell fate. Centriolar distal appendage protein ANKRD26 is the centrosomal PIDD1 receptor.The centrosome recruits the PIDD1 precursor, which undergoes subsequent autoproteolysis.Clustering of PIDD1‐positive extra‐centrosomes is required for PIDDosome activation.P53‐dependent PIDD1 transactivation leads to centrosome‐dependent PIDDosome activation even in the absence of extra‐centrosomes. [ABSTRACT FROM AUTHOR]

The PIDDosome is a Caspase-2-activating platform assembling in response to centrosome amplification or genotoxic stress. We have recently shown that both stimuli depend on ANKRD26 (ankyrin repeat domain-containing protein 26)-mediated localization of PIDD1 (p53-inducible protein with death domain) at the centrosome, demonstrating how this organelle can directly influence cell fate. [ABSTRACT FROM AUTHOR]

Interfering with mitosis for cancer treatment is an old concept that has proven highly successful in the clinics. Microtubule poisons are used to treat patients with different types of blood or solid cancer since more than 20 years, but how these drugs achieve clinical response is still unclear. Arresting cells in mitosis can promote their demise, at least in a petri dish. Yet, at the molecular level, this type of cell death is poorly defined and cancer cells often find ways to escape. The signaling pathways activated can lead to mitotic slippage, cell death, or senescence. Therefore, any attempt to unravel the mechanistic action of microtubule poisons will have to investigate aspects of cell cycle control, cell death initiation in mitosis and after slippage, at single-cell resolution. Here, we discuss possible mechanisms and signaling pathways controlling cell death in mitosis or after escape from mitotic arrest, as well as secondary consequences of mitotic errors, particularly sterile inflammation, and finally address the question how clinical efficacy of anti-mitotic drugs may come about and could be improved. [ABSTRACT FROM AUTHOR]

The PIDDosome is often used as the alias for a multi-protein complex that includes the p53-induced death domain protein 1 (PIDD1), the bipartite linker protein CRADD (also known as RAIDD) and the proform of an endopeptidase belonging to the caspase family, i.e. caspase-2. Yet, PIDD1 variants can also interact with a number of other proteins that include RIPK1 (also known as RIP1) and IKBKG (also known as NEMO), PCNA and RFC5, as well as nucleolar components such as NPM1 or NCL. This promiscuity in protein binding is facilitated mainly by autoprocessing of the full-length protein into various fragments that contain different structural domains. As a result, multiple responses can be mediated by protein complexes that contain a PIDD1 domain. This suggests that PIDD1 acts as an integrator for multiple types of stress that need instant attention. Examples are various types of DNA lesion but also the presence of extra centrosomes that can foster aneuploidy and, ultimately, promote DNA damage. Here, we review the role of PIDD1 in response to DNA damage and also highlight novel functions of PIDD1, such as in centrosome surveillance and scheduled polyploidisation as part of a cellular differentiation program during organogenesis. [ABSTRACT FROM AUTHOR]

Centrosomes, the main microtubule-organizing centers in animal cells, are replicated exactly once during the cell division cycle to form the poles of the mitotic spindle. Supernumerary centrosomes can lead to aberrant cell division and have been causally linked to chromosomal instability and cancer. Here,we report that an increase in the number of mature centrosomes, generated by disrupting cytokinesis or forcing centrosome overduplication, triggers the activation of the PIDDosome multiprotein complex, leading to Caspase-2-mediated MDM2 cleavage, p53 stabilization, and p21-dependent cell cycle arrest. This pathway also restrains the extent of developmentally scheduled polyploidization by regulating p53 levels in hepatocytes during liver organogenesis. Taken together, the PIDDosome acts as a first barrier, engaging p53 to halt the proliferation of cells carrying more than one mature centrosome to maintain genome integrity. [ABSTRACT FROM AUTHOR]

This is a correspondence about “Beclin‐1 is required for chromosome congression and proper outer kinetochore assembly”. [ABSTRACT FROM AUTHOR]

Both subunits of αβ-tubulin that comprise the core components of microtubules bind GTP. GTP binding to α-tubulin has a structural role, whereas β-tubulin binds and hydrolyses GTP to regulate microtubule dynamics. γ-tubulin, another member of the tubulin superfamily that seeds microtubule nucleation at microtubule-organizing centres, also binds GTP; however, the importance of this association remains elusive. To address the role of GTP binding to γ-tubulin, we systematically mutagenized the GTP contact residues in the yeast γ-tubulin Tub4. Tub4GTP-mutant proteins that exhibited greatly reduced GTP affinity still assembled into the small γ-tubulin complex. However, tub4GTP mutants were no longer viable, and had defects in interaction between γ-tubulin and αβ-tubulin, decreased microtubule nucleation and defects in microtubule organization. In vitro and in vivo data show that only γ-tubulin loaded with GTP nucleates microtubules. Our results suggest that GTP recruitment to γ-tubulin enhances its interaction with αβ-tubulin similarly to GTP recruitment to β-tubulin. [ABSTRACT FROM AUTHOR]

The article presents information on possible modes of action, proposed functions and regulation of caspase-2, a member of caspase family that are involved in cell death, inflammation and differentiation. It mentions that after activation, Caspase-2 is able to translocate from cytosol into the nucleus and confirms the hypothesis that it might act as a nuclear protease which orchestrate the response to DNA damage.

The spindle assembly checkpoint (SAC) restrains anaphase until all chromosomes become bi-oriented on the mitotic spindle. The SAC protein Mad2 can fold into two distinct conformers, open (O) and closed (C), and can asymmetrically dimerize. Here, we describe a monoclonal antibody that specifically recognizes the dimerization interface of C-Mad2. This antibody revealed several conformation-specific features of Mad2 in human cells. Notably, we show that Mad2 requires association with Mad1 to adopt the closed conformation and that the activity of the Mad1:C-Mad2 complex undergoes regulation by p31comet-dependent 'capping'. Furthermore, C-Mad2 antibody microinjection caused an abrupt termination of the SAC and accelerated mitotic progression. Remarkably, microinjection of a Mad1-neutralizing antibody triggered a comparable mitotic acceleration. Our study provides direct in vivo evidence for the model that a kinetochore complex of Mad1:C-Mad2 acts as a template to sustain the SAC and it challenges the distinction between SAC and mitotic timer. [ABSTRACT FROM AUTHOR]

Mitotic spindle formation and chromosome segregation depend critically on kinetochore--microtubule (KT-MT) interactions. A new protein, termed Spindly in Drosophila and SPDL-1 in C. elegans, was recently shown to regulate KT localization of dynein, but depletion phenotypes revealed striking differences, suggesting evolutionarily diverse roles of mitotic dynein. By characterizing the function of Spindly in human cells, we identify specific functions for KT dynein. We show that localization of human Spindly (hSpindly) to KTs is controlled by the Rod/Zw10/Zwilch (RZZ) complex and Aurora B. hSpindly depletion results in reduced inter-KT tension, unstable KT fibers, an extensive prometaphase delay, and severe chromosome misalignment. Moreover, depletion of hSpindly induces a striking spindle rotation, which can be rescued by co-depletion of dynein. However, in contrast to Drosophila, hSpindly depletion does not abolish the removal of MAD2 and ZW10 from KTs. Collectively, our data reveal hSpindly-mediated dynein functions and highlight a critical role of KT dynein in spindle orientation. [ABSTRACT FROM AUTHOR]

The performance of a narrow buffer strip in abating dissolved P, electrical conductivity and herbicides (terbuthylazine, alachlor, nicosulfuron, pendimethalin, linuron) in subsurface water coming from cropland was tested in an experiment carried out on the low plains of the Veneto Region (NE Italy). The experiment lasted from December 1997 to June 1999, monitoring subsurface water quality entering and exiting a buffer composed of a grass strip (5 m wide) and 1 m wide row of trees. Dissolved phosphorus concentrations were reduced by almost 100% passing through the buffer and in most cases exiting water satisfied the limit for avoiding eutrophication. A positive effect was also detected on ECW (reduced by 20%), while pH was not significantly altered. Herbicide concentration abatement varied between 60 and 90%, depending on the chemical and the time elapsed since application. [Copyright &y& Elsevier]

According to current belief, the molecular networks orchestrating cell death or exit from mitosis upon extended mitotic arrest do not interact, stubbornly executing two parallel biological programs and competing to define a stochastic decision between death and a chance for survival with uncertain destiny. However, recent findings by Diaz-Martinez et al () in this issue of The EMBO Journal now call for a reassessment of the 'competing network' hypothesis. [ABSTRACT FROM AUTHOR]

The metabolites of pesticides can contaminate groundwater and pose arisk to human health when this water is used for drinking. This paper reports the results of a laboratory study on aldicarb and its main metabolites, aldicarb sulfone and aldicarb sulfoxide. Aldicarb and its metabolites showed Koc values (6-31) which were lower than that of atrazine (55), indicating that they are very mobile in soil. They are less persistent than atrazine (DT50 = 25 days), with DT50 values from less than 1 day and up to 12 days.Aldicarb behaved as a non-leacher, whereas its metabolites clearly showed the characteristics of leachers. Aged residue leaching experiments showed that aldicarb can occur at high concentrations in the leachate, together with its two metabolites. The leachate composition depends on the incubation time of the parent compound. Aldicarb and its metabolites can form various mixtures in groundwater on the basis of the time elapsing between the application of the insecticide and the first significant rainfall. This study confirms the characteristics of contaminants of aldicarb and especially its metabolites, as reported in the literature. [ABSTRACT FROM AUTHOR]

Summary Receptor-interacting protein kinase (RIPK) 1 functions as a key mediator of tissue homeostasis via formation of Caspase-8 activating ripoptosome complexes, positively and negatively regulating apoptosis, necroptosis, and inflammation. Here, we report an unanticipated cell-death- and inflammation-independent function of RIPK1 and Caspase-8, promoting faithful chromosome alignment in mitosis and thereby ensuring genome stability. We find that ripoptosome complexes progressively form as cells enter mitosis, peaking at metaphase and disassembling as cells exit mitosis. Genetic deletion and mitosis-specific inhibition of Ripk1 or Caspase-8 results in chromosome alignment defects independently of MLKL. We found that Polo-like kinase 1 (PLK1) is recruited into mitotic ripoptosomes, where PLK1's activity is controlled via RIPK1-dependent recruitment and Caspase-8-mediated cleavage. A fine balance of ripoptosome assembly is required as deregulated ripoptosome activity modulates PLK1-dependent phosphorylation of downstream effectors, such as BUBR1. Our data suggest that ripoptosome-mediated regulation of PLK1 contributes to faithful chromosome segregation during mitosis. Graphical Abstract Highlights • A RIPK1/FADD/Caspase-8/cFLIP complex (ripoptosome) forms during mitosis • The mitotic kinase PLK1 binds to RIPK1 and is recruited into mitotic ripoptosomes • PLK1 is cleaved by Caspase-8 in a RIPK1-dependent manner • Deletion and inhibition of RIPK1 or Caspase-8 lead to chromosomal instability Liccardi et al. show that ripoptosome complexes assemble during physiological mitosis and regulate PLK1 activity via RIPK1-dependent recruitment and Caspase-8-mediated cleavage. These results reveal a direct role for RIPK1 and Caspase-8 in controlling chromosome segregation during mitosis. [ABSTRACT FROM AUTHOR]