A Kinase-Phosphatase Network that Regulates Kinetochore-Microtubule Attachments and the SAC

The KMN network (for KNL1, MIS12 and NDC80 complexes) is a hub for signalling at the outer kinetochore. It integrates the activities of two kinases (MPS1 and Aurora B) and two phosphatases (PP1 and PP2A-B56) to regulate kinetochore-microtubule attachments and the spindle assembly checkpoint (SAC). We will first discuss each of these enzymes separately, to describe how they are regulated at kinetochores and why this is important for their primary function in controlling either microtubule attachments or the SAC. We will then discuss why inhibiting any one of them individually produces secondary effects on all the others. This cross-talk may help to explain why all enzymes have been linked to both processes, even though the direct evidence suggests they each control only one. This chapter therefore describes how a network of kinases and phosphatases work together to regulate two key mitotic processes.</p

Causal assessment of smoking and tooth loss: A systematic review of observational studies

<p>Abstract</p> <p>Background</p> <p>Tooth loss impairs oral function. The aim of the present review was to evaluate the causal association between smoking and tooth loss on the basis of high-quality studies.</p> <p>Methods</p> <p>Relevant literature was searched and screened, and the methodological quality was assessed. Information on the strength of the association between smoking and tooth loss, the dose-response relationship and natural experimental data was collected and evaluated with respect to consistency and study design.</p> <p>Results</p> <p>Our literature search yielded 496 citations, and 6 cross-sectional and 2 cohort high-quality studies examining 58,755 subjects in four countries. All studies reported significant associations, although the strength of the association was usually moderate. Four studies reported dose-response relationships between exposure to smoking and the risk of developing tooth loss. A decrease in the risk of tooth loss for former smokers was evident in six studies. Interpretation of evidence for each element was consistent, despite some shortcomings regarding study type and population.</p> <p>Conclusions</p> <p>Based on the consistent evidence found with the existing biological plausibility, a causal association between smoking and tooth loss is highly likely. Further studies using a cohort design and different populations are necessary to confirm this association.</p

The Aurora Kinase in Trypanosoma brucei Plays Distinctive Roles in Metaphase-Anaphase Transition and Cytokinetic Initiation

Aurora B kinase is an essential regulator of chromosome segregation with the action well characterized in eukaryotes. It is also implicated in cytokinesis, but the detailed mechanism remains less clear, partly due to the difficulty in separating the latter from the former function in a growing cell. A chemical genetic approach with an inhibitor of the enzyme added to a synchronized cell population at different stages of the cell cycle would probably solve this problem. In the deeply branched parasitic protozoan Trypanosoma brucei, an Aurora B homolog, TbAUK1, was found to control both chromosome segregation and cytokinetic initiation by evidence from RNAi and dominant negative mutation. To clearly separate these two functions, VX-680, an inhibitor of TbAUK1, was added to a synchronized T. brucei procyclic cell population at different cell cycle stages. The unique trans-localization pattern of the chromosomal passenger complex (CPC), consisting of TbAUK1 and two novel proteins TbCPC1 and TbCPC2, was monitored during mitosis and cytokinesis by following the migration of the proteins tagged with enhanced yellow fluorescence protein in live cells with time-lapse video microscopy. Inhibition of TbAUK1 function in S-phase, prophase or metaphase invariably arrests the cells in the metaphase, suggesting an action of TbAUK1 in promoting metaphase-anaphase transition. TbAUK1 inhibition in anaphase does not affect mitotic exit, but prevents trans-localization of the CPC from the spindle midzone to the anterior tip of the new flagellum attachment zone for cytokinetic initiation. The CPC in the midzone is dispersed back to the two segregated nuclei, while cytokinesis is inhibited. In and beyond telophase, TbAUK1 inhibition has no effect on the progression of cytokinesis or the subsequent G1, S and G2 phases until a new metaphase is attained. There are thus two clearly distinct points of TbAUK1 action in T. brucei: the metaphase-anaphase transition and cytokinetic initiation. This is the first time to our knowledge that the dual functions of an Aurora B homolog is dissected and separated into two clearly distinct time frames in a cell cycle

A Conserved Role for SNX9-Family Members in the Regulation of Phagosome Maturation during Engulfment of Apoptotic Cells

Clearance of apoptotic cells is of key importance during development, tissue homeostasis and wound healing in multi-cellular animals. Genetic studies in the nematode Caenorhabditis elegans have identified a set of genes involved in the early steps of cell clearance, in particular the recognition and internalization of apoptotic cells. A pathway that orchestrates the maturation of phagosomes containing ingested apoptotic cells in the worm has recently been described. However, many steps in this pathway remain elusive. Here we show that the C. elegans SNX9-family member LST-4 (lateral signaling target) and its closest mammalian orthologue SNX33 play an evolutionary conserved role during apoptotic cell corpse clearance. In lst-4 deficient worms, internalized apoptotic cells accumulated within non-acidified, DYN-1-positive but RAB-5-negative phagosomes. Genetically, we show that LST-4 functions at the same step as DYN-1 during corpse removal, upstream of the GTPase RAB-5. We further show that mammalian SNX33 rescue C. elegans lst-4 mutants and that overexpression of truncated SNX33 fragments interfered with phagosome maturation in a mammalian cell system. Taken together, our genetic and cell biological analyses suggest that LST-4 is recruited through a combined activity of DYN-1 and VPS-34 to the early phagosome membrane, where it cooperates with DYN-1 to promote recruitment/retention of RAB-5 on the early phagosomal membrane during cell corpse clearance. The functional conservation between LST-4 and SNX33 indicate that these early steps of apoptotic phagosome maturation are likely conserved through evolution

Ybp2 Associates with the Central Kinetochore of Saccharomyces cerevisiae and Mediates Proper Mitotic Progression

The spindle checkpoint ensures the accurate segregation of chromosomes by monitoring the status of kinetochore attachment to microtubules. Simultaneous mutations in one of several kinetochore and cohesion genes and a spindle checkpoint gene cause a synthetic-lethal or synthetic-sick phenotype. A synthetic genetic array (SGA) analysis using a mad2Δ query mutant strain of yeast identified YBP2, a gene whose product shares sequence similarity with the product of YBP1, which is required for H2O2-induced oxidation of the transcription factor Yap1. ybp2Δ was sensitive to benomyl and accumulated at the mitotic stage of the cell cycle. Ybp2 physically associates with proteins of the COMA complex (Ctf19, Okp1, Mcm21, and Ame1) and 3 components of the Ndc80 complex (Ndc80, Nuf2, and Spc25 but not Spc24) in the central kinetochore and with Cse4 (the centromeric histone and CENP-A homolog). Chromatin-immunoprecipitation analyses revealed that Ybp2 associates specifically with CEN DNA. Furthermore, ybp2Δ showed synthetic-sick interactions with mutants of the genes that encode the COMA complex components. Ybp2 seems to be part of a macromolecular kinetochore complex and appears to contribute to the proper associations among the central kinetochore subcomplexes and the kinetochore-specific nucleosome

Spectrin-based skeleton as an actor in cell signaling

This review focuses on the recent advances in functions of spectrins in non-erythroid cells. We discuss new data concerning the commonly known role of the spectrin-based skeleton in control of membrane organization, stability and shape, and tethering protein mosaics to the cellular motors and to all major filament systems. Particular effort has been undertaken to highlight recent advances linking spectrin to cell signaling phenomena and its participation in signal transduction pathways in many cell types

Antitumor activity of a small-molecule inhibitor of the histone kinase Haspin

The approval of histone deacetylase inhibitors for treatment of lymphoma subtypes has positioned histone modifications as potential targets for the development of new classes of anticancer drugs. Histones also undergo phosphorylation events, and Haspin is a protein kinase the only known target of which is phosphorylation of histone H3 at Thr3 residue (H3T3ph), which is necessary for mitosis progression. Mitotic kinases can be blocked by small drugs and several clinical trials are underway with these agents. As occurs with Aurora kinase inhibitors, Haspin might be an optimal candidate for the pharmacological development of these compounds. A high-throughput screening for Haspin inhibitors identified the CHR-6494 compound as being one promising such agent. We demonstrate that CHR-6494 reduces H3T3ph levels in a dose-dependent manner and causes a mitotic catastrophe characterized by metaphase misalignment, spindle abnormalities and centrosome amplification. From the cellular standpoint, the identified small-molecule Haspin inhibitor causes arrest in G2/M and subsequently apoptosis. Importantly, ex vivo assays also demonstrate its anti-angiogenetic features; in vivo, it shows antitumor potential in xenografted nude mice without any observed toxicity. Thus, CHR-6494 is a first-in-class Haspin inhibitor with a wide spectrum of anticancer effects that merits further preclinical research as a new member of the family of mitotic kinase inhibitors

Polo kinase recruitment via the constitutive centromere-associated network at the kinetochore elevates centromeric RNA

The kinetochore, a multi-protein complex assembled on centromeres, is essential to segregate chromosomes during cell division. Deficiencies in kinetochore function can lead to chromosomal instability and aneuploidy-a hallmark of cancer cells. Kinetochore function is controlled by recruitment of regulatory proteins, many of which have been documented, however their function often remains uncharacterized and many are yet to be identified. To identify candidates of kinetochore regulation we used a proteome-wide protein association strategy in budding yeast and detected many proteins that are involved in post-translational modifications such as kinases, phosphatases and histone modifiers. We focused on the Polo-like kinase, Cdc5, and interrogated which cellular components were sensitive to constitutive Cdc5 localization. The kinetochore is particularly sensitive to constitutive Cdc5 kinase activity. Targeting Cdc5 to different kinetochore subcomplexes produced diverse phenotypes, consistent with multiple distinct functions at the kinetochore. We show that targeting Cdc5 to the inner kinetochore, the constitutive centromere-associated network (CCAN), increases the levels of centromeric RNA via an SPT4 dependent mechanism